Patent Publication Number: US-2021163900-A1

Title: Nucleic acid modification with tools from oxytricha

Description:
CROSS REFERENCE TO RELATED APPLICATIONS 
     The present application is a continuation of PCT international application no. PCT/US2019/042625, filed on Jul. 19, 2019, which claims benefit of claims benefit of U.S. Provisional patent Application Ser. No. 62/701,536, filed on Jul. 20, 2018, and U.S. Provisional patent Application Ser. No. 62/848,414, filed on May 15, 2019. The entire contents of the aforementioned applications are incorporated by reference as if recited in full herein. 
    
    
     GOVERNMENT FUNDING 
     This invention was made with government support under GM059708 and GM122555 awarded by the National Institutes of Health. The government has certain rights in the invention. 
    
    
     FIELD OF DISCLOSURE 
     The present disclosure provides, inter alia, various methods, kits and compositions for modifying nucleic acid using MTA1c or any components thereof. Such embodiments may be used to treat disease and as research tools. 
     INCORPORATION BY REFERENCE OF SEQUENCE LISTING 
     This application contains references to amino acids and/or nucleic acid sequences that have been filed concurrently herewith as sequence listing text file “CU19015-PCT-seq.txt”, file size of 478 KB, created on Aug. 28, 2019. The aforementioned sequence listing is hereby incorporated by reference in its entirety. 
     BACKGROUND OF THE DISCLOSURE 
     Covalent modifications on DNA have long been recognized as a hallmark of epigenetic regulation. DNA N6-methyladenine (6 mA) has recently come under scrutiny in eukaryotic systems, with proposed roles in retrotransposon or gene regulation, transgenerational epigenetic inheritance, and chromatin organization (Luo et al., 2015). 6 mA exists at low levels in  Arabidopsis thaliana  (0.006%-0.138% 6 mA/dA), rice (0.2%),  C. elegans  (0.01%-0.4%),  Drosophila  (0.001%-0.07%),  Xenopus laevis  (0.00009%), mouse embryonic stem cells (ESCs) (0.0006-0.007%), human cells (Greer et al., 2015; Koziol et al., 2016; Liang et al., 2018; Wu et al., 2016; Xiao et al., 2018; Zhang et al., 2015; Zhou et al., 2018), and the mouse brain (Yao et al., 2017), although it accumulates in abundance (0.1%-0.2%) during vertebrate embryogenesis (Liu et al., 2016). Disruption of DMAD, a 6 mA demethylase, in the  Drosophila  brain leads to the accumulation of 6 mA and Polycomb-mediated silencing (Yao et al., 2018). The existence of 6 mA in mammals remains a subject of debate. Quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis of HeLa and mouse ESCs failed to detect 6 mA above background levels (Schiffers et al., 2017). A recent study, however, reported that loss of 6 mA in human cells promotes tumor formation (Xiao et al., 2018), suggesting that 6 mA is a biologically relevant epigenetic mark. 
     In contrast to metazoa, 6 mA is abundant in various unicellular eukaryotes, including ciliates (0.18%-2.5%) (Ammermann et al., 1981; Cummings et al., 1974; Gorovsky et al., 1973; Rae and Spear, 1978), and the green algae  Chlamydomonas  (0.3%-0.5%) (Fu et al., 2015; Hattman et al., 1978). High levels of 6 mA (up to 2.8%) were also recently reported in basal fungi (Mondo et al., 2017). Ciliates have long served as powerful models for the study of chromatin modifications (Brownell et al., 1996; Liu et al., 2007; Strahl et al., 1999; Taverna et al., 2002; Wei et al., 1998). They possess two structurally and functionally distinct nuclei within each cell (Bracht et al., 2013; Yerlici and Landweber, 2014). In the ciliate  Oxytricha trifallax , the germline micronucleus is transcriptionally silent and contains ˜100 megabase-sized chromosomes (Chen et al., 2014). In contrast, the somatic macronucleus is transcriptionally active, being the sole locus of Pol II-dependent RNA production in non-developing cells (Khurana et al., 2014). The  Oxytricha  macronuclear genome is extraordinarily fragmented, consisting of ˜16,000 unique chromosomes with a mean length of ˜3.2 kb, most encoding a single gene. Macronuclear chromatin yields a characteristic ˜200 bp ladder upon digestion with micrococcal nuclease, indicative of regularly spaced nucleosomes (Gottschling and Cech, 1984; Lawn et al., 1978; Wada and Spear, 1980). Yet it remains unknown how and where nucleosomes are organized within these miniature chromosomes and if this in turn regulates (or is regulated by) 6 mA deposition. 
     SUMMARY OF THE DISCLOSURE 
     The ciliate  Oxytricha  is a natural source of tools for RNA-guided genome reorganization and other nucleic acid modification. Long template RNAs instruct new linkages between pieces of DNA (Nowacki et al. 2008), and small RNAs instruct which DNA segments to keep (Fang et al. 2012) or eliminate. Foreseeable uses of these or other machinery derived from the  Oxytricha  genome include in vitro and/or in vivo modification of nucleic acids. 
     Intriguingly, in green algae, basal yeast, and ciliates, 6 mA is enriched in ApT dinucleotide motifs within nucleosome linker regions near promoters (Fu et al., 2015; Hattman et al., 1978; Karrer and VanNuland, 1999; Mondo et al., 2017; Pratt and Hattman, 1981; Wang et al., 2017). In the present disclosure, four ciliate proteins-named MTA1, MTA9, p1, and p2—have been identified as being necessary for 6 mA methylation in a complex form termed MTA1c. MTA1 and MTA9 contain divergent MT-A70 domains, while p1 and p2 are homeobox-like proteins that likely function in DNA binding. The present disclosure delineates key biochemical properties of this methyltransferase and dissects the function of 6 mA in vitro and in vivo. 
     The present disclosure provides a novel ciliate enzyme “MTA1” effective for N6-methyladenine (m6dA) methylation of DNA (see, e.g., Appendix 4). MTA1 has been identified in a ciliate,  Tetrahymena thermophila , and its functional role validated in m6dA methylation in  Oxytricha . (See, Genbank ID: XP 001032074.3 [ Tetrahymena  MTA1] and EJY79437.1 [ Oxytricha  MTA1]). MTA1 is evolutionarily distinct from all known m6dA methyltransferases. Evolutionary analysis reveals that it is present in ciliates (including  Oxytricha  and  Tetrahymena ), algae, and basal fungi, but not multicellular eukaryotes. MTA1 exhibits a unique substrate specificity in vivo, being essential for the deposition of dimethylated AT (5′-A*T-3′/3′-TA*-5′), as well as a wide range of other motifs in vivo ( FIGS. 1A-1B ). The inventors have been actively characterizing the biochemical properties and enzymology of  Tetrahymena  and  Oxytricha  MTA1, including its binding partners, in vitro substrate specificity (DNA vs. RNA and sequence motifs therein), methylation kinetics, and structural basis of these activities. 
     The present disclosure provides that MTA1c or any components thereof presents immediate commercial applications in: 1) generation of DNA substrates containing m6dA at locations distinct from known m6dA methyltransferases, circumventing the need for slow, expensive synthesis of methylated DNA; and 2) rational design of N6-adenine methylating enzymes with novel substrate specificities. 
     Accordingly, one embodiment of the present disclosure is a method of modifying a nucleic acid from a cell, the cell derived from a multicellular eukaryote. This method comprises the steps of: (a) obtaining the nucleic acid from the cell; and (b) contacting the nucleic acid with MTA1c or any components thereof under conditions effective to methylate the nucleic acid. 
     The modified base, m6dA, has been discovered in a wide range of eukaryotes, including humans. m6dA levels are significantly reduced in gastric and liver cancer tissues, and disruption of m6dA promotes tumor formation (Xiao et al. 2018). As disclosed herein, MTA1 is a novel m6dA “writer”, paving the way for cost-effective methods to understand mechanisms of m6dA function in biomedically relevant models. 
     Accordingly, another embodiment of the present disclosure is a method of treating or ameliorating the effects of a disease characterized by an abnormal level of m6dA in a subject. This method comprises administering to the subject an amount of MTA1c or any components thereof effective to modulate m6dA levels in the subject. In some embodiments, the modulation comprises restoring m6dA levels to normal or near-normal ranges in the subject. 
     Another embodiment of the present disclosure is a pharmaceutical composition comprising MTA1c or any components thereof that is effective to modulate m6dA levels in a subject in need thereof and a pharmaceutically acceptable carrier, diluent, adjuvant or vehicle. 
     Yet another embodiment of the present disclosure is a kit for treating or ameliorating the effects of a disease characterized by an abnormal level of m6dA in a subject, such as, e.g., cancer, comprising an effective amount of MTA1c or any components thereof, packaged together with instructions for its use. 
     Another embodiment of the present disclosure is a cell line obtained from a multicellular eukaryote comprising a nucleic acid encoding MTA1c or any components thereof and/or an MTA1c protein complex or any components thereof. As used herein, a “cell line” refers to all types of cell lines such as, e.g., immortalized cell lines and primary cell lines. In certain embodiments, the nucleic acid encoding MTA1c or any components thereof is operably linked to a recombinant expression vector. 
     Another embodiment of the present disclosure is a recombinant expression vector comprising a polynucleotide encoding MTA1c or any components thereof. 
     Still another embodiment of the present disclosure is a transgenic organism whose genome comprises a transgene comprising a nucleotide sequence encoding MTA1c or any components thereof. Non-limiting examples of possible organism include an archaea, a bacterium, a eukaryotic single-cell organism, algae, a plant, an animal, an invertebrate, a fly, a worm, a cnidarian, a vertebrate, a fish, a frog, a bird, a mammal, an ungulate, a rodent, a rat, a mouse, and a non-human primate. 
     The present disclosure also provides a method of identifying protein binding sites on DNA. This method comprises the steps of: (a) providing DNA; (b) contacting the DNA with MTA1c or any components thereof under conditions effective to methylate the DNA; (c) contacting the DNA with one or more proteins; (d) contacting the DNA with an enzyme effective to hydrolize the DNA in positions where no protein binding occurs; (e) removing the DNA bound protein; and (f) isolating and sequencing the DNA fragments. In certain embodiments, the one or more proteins in step (c) comprise histone octamers. 
     Another embodiment of the present disclosure is a method of mediating DNA N6-adenine methylation. This method comprises the steps of: (a) providing DNA; and (b) contacting the DNA with MTA1c or any components thereof under conditions effective to methylate the DNA. 
     Another embodiment of the present disclosure is a method of modulating nucleosome organization and/or transcription in a cell, comprising providing to the cell an agent that is effective to modulate the expression of MTA1c or any components thereof. 
     The present disclosure also provides a method of generating a synthetic chromosome. This method comprises the steps of: (a) generating chromosome segments containing terminal restriction sites, wherein the chromosome segments comprise one or more m6dA bases; (b) digesting the chromosome segments with a restriction enzyme; and (c) purifying and ligating the digested chromosome segments to form a synthetic chromosome. In some embodiments, the method further comprises enriching the synthetic chromosome. A synthetic chromosome made by the method above is also provided. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
       The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee. 
       The following drawings form part of the present specification and are included to further demonstrate certain aspects of the present disclosure. The disclosure may be better understood by reference to one or more of these drawings in combination with the detailed description of specific embodiments presented herein. 
         FIGS. 1A-1E  show epigenomic profiles of  Oxytricha  chromosomes. 
         FIG. 1A  shows meta-chromosome plots of chromatin organization at  Oxytricha  macronuclear chromosome ends. Heterodimeric telomere end-binding protein complexes (orange ovals) protect each end in vivo. Horizontal red bar: promoter. The 5′ chromosome end is proximal to TSSs. Nucleosome occupancy, normalized Mnaseseq coverage; 6 mA, total 6 mA number; Transcription start sites, total number of called TSSs. 
         FIG. 1B  shows histograms of the total number of 6 mA marks within each linker in  Oxytricha  chromosomes. Distinct linkers are depicted as horizontal blue lines. 
         FIG. 1C  shows that poly(A)-enriched RNA-seq levels positively correlate with 6 mA. Genes are sorted according to the total number of 6 mA marks 0-800 bp downstream of the TSS. FPKM, fragments per kilobase of transcript per million mapped RNA-seq reads. Notch in the boxplot denotes median, ends of boxplot denote first and third quartiles, upper whisker denotes third quantile+1.5× interquartile range (IQR), and lower whisker denotes data quartile 1-1.5×IQR. 
         FIG. 1D  shows that composite analysis of 65,107 methylation sites reveals that 6 mA (marked with 1 occurs within a 5′-ApT-3′ dinucleotide motif. 
         FIG. 1E  provides the distribution of various 6 mA dinucleotide motifs across the genome. Asterisk, 6 mA. 
         FIGS. 2A-2G  show purification and characterization of the ciliate 6 mA methyltransferase. 
         FIG. 2A  provides phylogenetic analysis of MT-A70 proteins. Bold MTA1 and MTA9 genes are experimentally characterized in this study. Paralogs of MTA1 and MTA9 are labeled as “-B.” Posterior probabilities &gt;0.65 are shown. Gray triangle represents outgroup of bacterial sequences. The complete phylogenetic tree is shown in  FIG. 9G . Gene names are in Table 5. Tth,  Tetrahymena thermophila ; Otri,  Oxytricha trifallax.    
         FIG. 2B  shows the phylogenetic distribution of the occurrence of ApT 6 mA motifs and MT-A70 protein families. Filled square denotes its presence in a taxon. The basal yeast clade is comprised of  L. transversale, A. repens, H. vesiculosa, S. racemosum, L. pennispora, B. meristosporus, P. finnis , and  A. robustus.    
         FIG. 2C  is an experimental scheme depicting the partial purification of DNA methyltransferase activity from  Tetrahymena  nuclear extracts. 
         FIG. 2D  show gene expression and protein abundance of candidate genes in partially purified  Tetrahymena  nuclear extracts. UniProt IDs are listed in Table 5. RNA-seq data are from (Xiong et al. 2012). FPKM, fragments per kilobase of transcript per million mapped RNA-seq reads. Low, Mid, and High DNA methylase activity correspond to fractions eluting from the Nuvia cPrime and Superdex 200 columns in  FIG. 2C . Total spectrum counts, total number of LC-MS/MS fragmentation spectra that match peptides from a target protein. 
         FIG. 2E  shows DNA methyltransferase assay using [3H]SAM. Vertical axis represents scintillation counts. Error bars represent SEM (n=3). 
         FIG. 2F  shows dot blot assay using cold SAM. 
         FIG. 2G  shows DNA methyltransferase assay performed on different nucleic acid substrates in the presence of MTA1, MTA9, p1, and p2. Sense ssDNA are 5′→3′; antisense are 3′→5′. ApT dinucleotides are labeled in bold red. Horizontal blue lines in hemimethylated dsDNA substrates denote possible locations where 6 mA may be installed by EcoGII (prior to this assay). Relative activity denotes scintillation counts normalized against the unmethylated 27 bp dsDNA substrate with two ApT motifs (top-most dsDNA substrate). An enlarged bar plot of relative activity on 27 bp unmethylated dsDNA substrates is included in  FIG. 10K . Error bars represent SEM (n=3). 
         FIGS. 3A-3E  show genome-wide loss of 6 mA in mta1 mutants. 
         FIG. 3A  shows schematic depicting the disruption of  Oxytricha  MTA1 open reading frame. Flanking dark blue bars: 5′ and 3′ UTR; yellow, open reading frame; red, retention of 62 bp ectopic DNA segment; gray bar, intron; Internal light blue bar, annotated MT-A70 domain; ATG, start codon; TGA, stop codon. Agarose gel analysis shows PCR confirmation of ectopic DNA retention. 
         FIG. 3B  shows dot blot analysis of RNase-treated genomic DNA. 
         FIG. 3C  shows histogram of 6 mA counts near 5′ and 3 ′ Oxytricha  chromosome ends. Inset depicts histogram of fold change in total 6 mA in each chromosome, between mutant and wild-type cell lines. 
         FIG. 3D  shows that chromosomes are sorted into 10 groups according to total 6 mA in wild-type cells (blue boxplots). For each group, the total 6 mA per chromosome in mutants and the difference in total 6 mA per chromosome are plotted below. Boxplot features are as described in  FIG. 1C . 
         FIG. 3E  shows motif distribution in wild-type and mta1 mutants. Loss of ApT dimethylated motif is underlined. 
         FIGS. 4A-4E  show effects of 6ma on nucleosome organization in vitro and in vivo. 
         FIG. 4A  shows the experimental workflow for the generation of mini-genome DNA. 
         FIG. 4B  shows agarose gel analysis of  Oxytricha  gDNA (Native) and mini-genome DNA before chromatin assembly. 
         FIG. 4C  shows that methylated regions exhibit lower nucleosome occupancy in vitro but not in vivo. Overlapping 51 bp windows were analyzed across 98 chromosomes. For each window, the change in nucleosome occupancy in the absence versus presence of 6 mA was calculated. Boxplot features are as described in  FIG. 1C . p values were calculated using a two-sample unequal variance t test. N.S., non-significant, with p&gt;0.05. 
         FIG. 4D  shows the reduction in nucleosome occupancy at methylated loci in vitro (black arrowheads). For in vitro MNase-seq, +6 mA refers to chromatin assembled on  Oxytricha  gDNA, while −6 mA denotes chromatin assembled on mini-genome DNA. The vertical axis for SMRT-seq data denotes confidence score [−10 log(p value)] of detection of 6 mA, while that for in vitro MNase-seq data denotes nucleosome occupancy. 
         FIG. 4E  shows no change in nucleosome occupancy in linker regions despite loss of 6 mA in mta1 mutants. Vertical axes are the same as  FIG. 4D . 
         FIGS. 5A-5C  show modular synthesis of full-length  Oxytricha  chromosomes. 
         FIG. 5A  shows features of the chromosome selected for synthesis. Gray boxes represent exons. All data tracks represent normalized coverage except for SMRT-seq, which represents the confidence score [−10 log(p value)] of detection of each methylated base. 
         FIG. 5B  shows the schematic of chromosome construction. Different colors denote DNA building blocks ligated to form the full-length chromosome. Precise 6 mA sites (bold red) represent cognate 6 mA positions revealed by SMRT-seq in native genomic DNA. These are introduced via oligonucleotide synthesis. For chromosome 5, 6 mA sites (non-bold red) represent possible locations ectopically installed by a bacterial 6 mA methyltransferase, EcoGII. Intervening sequence within chromosomes 5 and 6 is represented as “ . . . ”. 
         FIG. 5C  shows native polyacrylamide gel analysis and anti-6 mA dot blot analysis of building blocks and purified synthetic chromosomes. 
         FIGS. 6A-6E  show quantitative modulation of nucleosome occupancy by 6 mA. 
         FIG. 6A  shows the experimental workflow. Chromatin is assembled using either salt dialysis or the NAP1 histone chaperone. Italicized blue steps are selectively included. 
         FIG. 6B  shows the tiling qPCR analysis of synthetic chromosome with cognate 6 mA sites. Horizontal gray box represents annotated gene, and vertical black lines depict native 6 mA positions. Horizontal blue bars span −100 bp regions amplified by qPCR. Red horizontal lines represent the region containing 6 mA. Hemi methyl chromosomes contain 6 mA on the antisense and sense strands, respectively, while the Full methyl chromosome has 6 mA on both strands. Black arrowheads: decrease in nucleosome occupancy specifically at the 6 mA cluster. 
         FIG. 6C  shows the tiling qPCR analysis of ectopically methylated synthetic chromosome. Vertical black lines illustrate possible 6 mA sites installed enzymatically. Red arrowheads: decrease in nucleosome occupancy in the ectopically methylated region. Black arrowheads: position of cognate 6 mA sites (not in this construct). 
         FIG. 6D  shows the tiling qPCR analysis of chromatin from  FIG. 6B  that is subsequently incubated with ACF and/or ATP. ACF equalizes nucleosome occupancy between the 6 mA cluster and flanking regions in the presence of ATP (black line). Nucleosome occupancy at the methylated region is not restored to the same level as the unmethylated control (black arrowheads). 
         FIG. 6E  shows that MNase-seq analysis of chromatin is assembled on native gDNA (“+” 6 mA) and mini-genome DNA (“−” 6 mA) using NAP1±ACF and ATP. p values were calculated using a two-sample unequal variance t test. 
         FIGS. 7A-7F  show effects of 6 mA on gene expression and cell viability in vivo. 
         FIG. 7A  shows the following: Horizontal axis: the mean RNA-seq counts across all biological replicates from wild-type and mta1 mutant data for each gene. Vertical axis: log 2(fold change) in gene expression (mutant/wild type). 
         FIG. 7B  shows that upregulated genes tend to be sparsely methylated compared to randomly subsampled genes (gray lines). 
         FIG. 7C  shows RNA-seq analysis of MTA1 expression during the sexual cycle of  Oxytricha . RNA-seq time course data are from Swart et al. (2013). The total duration of the sexual cycle is ˜60 h. 
         FIG. 7D  shows survival analysis of  Oxytricha  cells during the sexual cycle. The total cell number at each time point is normalized to 27 h data to obtain the percentage survival. Error bars represent SEM (n=4). 
         FIG. 7E  is a model illustrating the impact of 6 mA methylation by MTA1c on nucleosome organization and gene expression. 
         FIG. 7F  shows the comparison of DNA and RNA N6-adenine methyltransferases. Blue denotes catalytic subunit; yellow denotes subunit with predicted DNA or RNA binding domain. 
         FIGS. 8A-8B  show MS analysis of 6 mA in ciliate DNA. 
         FIG. 8A  shows that  Oxytricha  and  Tetrahymena  genomic DNA were digested into nucleosides using degradase enzyme mix, followed by analysis using reverse-phase HPLC and mass spectrometry. Isotopically labeled dA and 6 mA standards ( 15 N5-dA and D3-6 mA) were mixed with each sample to allow quantitative measurement of endogenous dA and 6 mA concentrations. MS/MS analysis of labeled dA and 6 mA standards confirmed the mass of the nucleobase. Fluted peaks with expected masses of dA and 6 mA, and with highly similar retention times (RT) to internal standards are detected in  Oxytricha  and  Tetrahymena  nucleosides. 
         FIG. 8B  shows the quantitation of dA and 6 mA levels in  Oxytricha  and  Tetrahymena  gDNA using internal isotopically labeled nucleoside standards. The detected level of 6 mA in  Tetrahymena  gDNA agrees with earlier reports (Gorovsky et al., 1973; Pratt and Hallman, 1981). The calculated abundance of 6 mA relative to (dA+6 mA) in  Oxytricha  is ˜0.71%, which is similar to the estimate from SMRT-seq base calls (0.78-1.04%). Note that the calculation from SMRT-seq data is expected to be an overestimate because 6 mA is scored at being present or absent at each site in the genome for this purpose. In actual fact, 6 mA sites may be partially methylated ( FIG. 11A ). Neither 6 mA nor dA was detected from LC-MS analysis of  Oxytricha  culture media, arguing against spurious signal arising from contamination or overall technical handling. The PacBio and LC-MS measurements of % 6 mA in  Oxytricha  are both similar to thin layer chromatography analysis of nucleotides (0.6-0.7%) from a distinct but closely related species,  Oxytricha fallax  (Rae and Spear, 1978). 
         FIGS. 9A-9K  show analysis of 6 mA and methyltransferase components in  Tetrahymena.    
         FIG. 9A  shows  Tetrahymena  MNase-seq data from (Beh et al., 2015), while SMRT-seq data were generated in the present disclosure. Meta-chromosome plots overlaying in vivo MNase-seq (nucleosome occupancy) and SMRT-seq (6 mA), relative to annotated transcription start sites. 6 mA lies mainly within nucleosome linker regions, between the +1, +2, +3, and +4 nucleosomes. 
         FIG. 9B  shows histograms of the total number of 6 mA marks within each linker in  Tetrahymena  genes. Calculations are performed as described in  FIG. 1B . Distinct linkers are highlighted with horizontal bold blue lines. 
         FIG. 9C  shows the relationship between transcriptional activity and total number of 6 mA marks in  Tetrahymena  genes. Analysis is performed as in  FIG. 1C . RNA-seq data was obtained from (Xiong et al., 2012). 
         FIG. 9D  shows that composite analysis of 441,618 methylation sites reveals that 6 mA occurs within a 5′-ApT-3′ dinucleotide motif in  Tetrahymena , consistent with previous experiments (Bromberg et al., 1982; Wang et al., 2017) and similar to  Oxytricha.    
         FIG. 9E  shows distribution of various 6 mA dinucleotide motifs across the genome. 
         FIG. 9F  shows organization of transcription (mRNA-seq), nucleosome organization (MNase-seq), and 6 mA (SMRT-seq) in a  Tetrahymena  gene. 
         FIG. 9G  shows that all sequences used for phylogeny construction are listed in Table 1. Abbreviations: Cel:  Caenorhabditis elegans ; Ath:  Arabidopsis thaliana ; Sra:  Syncephalastrum racemosum ; Hve:  Hesseltinella vesiculosa ; Are:  Absidia repens ; Dre:  Danio redo ; Has:  Homo sapiens ; Ssc:  Sus scrota ; Mmu:  Mus musculus ; Xla:  Xenopus laevis ; Dme:  Drosophila melanogaster ; Cre:  Chlamydomonas reinhardtii ; Ltr:  Lobosporangium transversale ; Lpe:  Linderina pennispora ; Bme:  Basidiobolus meristosporus ; Pfi:  Piromyces finnis ; Aro:  Anaeromyces robustus ; Tth:  Tetrahymena thermophila ; Otri:  Oxytricha trifallax . This Bayesian phylogenetic tree of MT-A70 proteins is the same as in  FIG. 2A , except that all sequences are now included and labeled. TAMT-1 proteins are named according to (Luo et al., 2018). 
         FIG. 9H  shows Bayesian phylogenetic tree of p1 proteins. 
         FIG. 9I  shows Bayesian phylogenetic tree of p2 proteins. Dashed box depicts outgroup consisting of vertebrate SNAPC4 genes. These genes bear weak similarity to the homeobox-like domain of p2 proteins, but do not group phylogenetically with them and are therefore unlikely to be functionally homologs. 
         FIG. 9J  shows phylogenetic distribution of ApT 6 mA motif and various proteins, as depicted in  FIG. 2B , but now also including TAMT-1, p1, and p2 proteins. Filled boxes denote the presence of a particular protein in a taxon. Open dashed boxes indicate the presence of SNAPC4 genes in vertebrates. 
         FIG. 9K  shows the gene expression profiles of  Tetrahymena  MTA1, MTA9, p1 and p2. Microarray counts represent poly(A)′ expression levels, and are obtained from TetraFGD (Miao et al., 2009; Xiong et al., 2011). MTA1, MTA9, p1 and p2 were found in our study to co-elute with 6 mA methylase activity. On the other hand, TAMT-1 is a putative DNA methyltransferase described by (Luo et al., 2018). The horizontal axis categories beginning with “S” and “C” represent the number of hours since the onset of starvation and conjugation (mating), respectively. “Low,” “Med,” and “High” denote relative cell densities during log-phase growth. Blue and orange traces represent data from two biological replicates. Green and red shaded regions show the peaks in poly(A)* RNA expression in vegetative growth and conjugation, respectively, for MTA1, MTA9, p1 and p2. Note that their expression pattern differs from TAMT-1. 
         FIGS. 10A-10N  show further characterization of 6 mA methyltransferase activity and MTA1c. 
         FIG. 10A  shows that fractionation of nuclear extracts on a Q Sepharose column results in two distinct peaks of DNA methyltransferase activity, denoted as “Low Salt sample” and “High Salt sample” by black horizontal bars. FT denotes column flow-through. The DNA methyltransferase assay is performed as in  FIG. 2E . The salt concentration at which individual fractions elute from the column is plotted against DNA methyltransferase activity of each fraction (counts per minute). Inset shows DNA methyltransferase activity of the input nuclear extract, flowthrough from the Q Sepharose column, and blank control (nuclear extract buffer). Orange and blue plots denote replicates derived from independent preparations of nuclear extract. 
         FIG. 10B  is DNA methyltransferase assay showing that the activity from nuclear extracts is heat-sensitive and requires addition of DNA and SAM. Error bars represent s.e.m. (n=3). 
         FIG. 10C  is dot blot showing that nuclear extracts mediate 6 mA methylation. Note that the low salt sample has substantial DNase activity, resulting in a lower amount of DNA available for dot blot analysis. DNA substrate, nuclear extract, and SAM cofactor were mixed as in panels A and B. The DNA was subsequently purified and used for dot blot analysis. 
         FIG. 10D  shows domain organization of  Tetrahymena  MTA1, MTA9, p1, and p2. Protein domains are predicted using hmmscan on the EMBL-EBI webserver (Finn et al., 2015). “aa” denotes amino acids. Start and end coordinates of each domain are stated below each polypeptide. 
         FIG. 10E  shows the sequence alignment of human (Hsa) METTL3 with  Tetrahymena  (Tth) and  Oxytricha  (Otri) MTA1/MTA9, within the MT-A70 domain. Horizontal black bars underscore the DPPW catalytic motif, and the N549/0550 residues in human METTL3 that interact with the ribose moiety of the SAM cofactor. Note that the DPPW catalytic motif is conserved in MTA1 but not MTA9. 
         FIG. 10F  shows dot blot analysis of hemimethylated dsDNA substrates. Sense or antisense oligonucleotides were first individually methylated using the EcoGII bacterial 6 mA methyltransferase. Each methylated ssDNA was subsequently purified and annealed with an unmethylated complementary strand to form hemimethylated constructs. 
         FIG. 10G  shows SDS-PAGE analysis of recombinant proteins. Full length proteins were expressed and purified from  E. coli . Bands of expected size are indicated with a black arrowhead. 
         FIG. 10H  is methyttransferase assay using radiolabeled SAM on DNA and RNA substrates, coupled with gel analysis of nucleic acid integrity. ssRNA and dsRNA were produced by in vitro transcription from the 350 bp dsDNA template using 17 RNA polymerase, and subsequently purified before use in this assay. Methyltransferase activity on equimolar amounts of each substrate was measured after incubation at 37° C. for 6 hr, and depicted as either scintillation counts (Counts per minute), or normalized to the 350 bp dsDNA sample (Relative activity). Only dsDNA, and not dsRNA or ssRNA, was methylated. Activity measurements are represented as scintillation counts (counts per minute). In addition, aliquots from each reaction containing DNA or RNA substrate and recombinant MTA1c (ie. MTA1, MTA7, p1 and p2 proteins) were withdrawn at 0, 1, 2, 3, or 6 hr during the 37° C. incubation, purified using phenol:chloroform extraction and ethanol precipitation, and subsequently analyzed on a non-denaturing agarose gel. Both dsDNA and dsRNA substrates remained intact after 6 hr. The ssRNA migrates more diffusely on a nondenaturing agarose gel, with some decrease in size over time, suggesting partial degradation and/or RNA folding; however, there is no detectable methylation of ssRNA despite a significant presence on the agarose gel after 6 hr at 37° C. It is unlikely that this species is too short to be methylated, since MTA1c can methylate significantly shorter substrates such as 27 bp dsDNA ( FIGS. 2G, 10I, 10J, and 10K ). Error bars represent s.e.m. (n=3). 
         FIG. 10I  is DNA methyltransferase assay using radiolabeled SAM, on ssDNA oligonucleotides or annealed dsDNA substrates. All four recombinant MTA1c protein components—MTA1, MTA9, p1, and p2—were included in each sample. Activity measurements are represented as scintillation counts (counts per minute). dsDNA substrates were prepared by annealing ssDNA oligonucleotides, as in  FIG. 2G . Sense ssDNA nucleotide sequences are depicted in the 5′ 3′ direction, while antisense ssDNA is depicted as 3′ 5′. Error bars represent s.e.m. (n=3). 
         FIG. 10J  is control [ 3 H]SAM assay using hemimethylated dsDNA. Reactions depicted in red represent hemimethylated dsDNA incubated with [3H]SAM in the absence of recombinant MTA1c (MTA1, MTA9, p1, and p2 proteins). These reactions showed no methyltransferase activity, verifying that there is no contaminating EcoGII methyltransferase in hemimethylated dsDNA preparations. Activity measurements are shown as scintillation counts, or as “Relative Activity” (normalized against the sample containing unmethylated DNA substrate, [3H]SAM, and MTA1c protein). Hemimethylated dsDNA substrates in this panel are the same as those used in  FIG. 2G . The unmethylated dsDNA substrate used in this panel is the same as the top-most dsDNA substrate in  FIG. 2G , with two uninterrupted ApT dinucleotides. Error bars represent s.e.m. (n=3). 
         FIG. 10K  is DNA methyltransferase assay using radiolabeled SAM, on dsDNA substrates with disrupted ApT dinucleotides. All four recombinant MTA1c protein components—MTA1, MTA9, p1, and p2—were included in each sample. Activity measurements are normalized against the parent dsDNA construct with two uninterrupted ApT dinucleotides (top-most construct in this panel). ApT dinucleotide positions are labeled in bold red. Note that the parent dsDNA construct is identical to that in  FIG. 10L . Error bars represent s.e.m. (n=3). 
         FIG. 10L  is DNA methyitransferase assay using radiolabeled SAM, on dsDNA substrates with shifted ApT dinucleotides. All four recombinant MTA1c protein components—MTA1, MTA9, p1, and p2—were included in each sample. Activity measurements are normalized against the parent dsDNA construct with two uninterrupted ApT dinucleotides (top-most construct in this panel). The parent construct is identical to that in  FIG. 10K . ApT dinucleotides are labeled in bold red. The adjacent nucleotides are labeled in bold black to highlight the 4-mer sequence that contains each ApT dinucleotide. Error bars represent s.e.m. (n=3). 
         FIG. 10M  shows motif frequencies of all 4-mer sequences containing methylated ApT dinucleotides in the  Tetrahymena  and  Oxytricha  genomes. A′ denotes 6 mA. The 4-mers TA′TA and CKTT are colored in red and blue, respectively, to highlight their large difference in genomic frequencies. 
         FIG. 10N  shows motif frequencies of 4-mer sequences—regardless of methylation state—in  Tetrahymena  and  Oxytricha . These were calculated from genomic sequence between the 5′ chromosome end and the +4 nucleosome peak ( Oxytricha ), or between the TSS and the +4 nucleosome peak ( Tetrahymena ). Analysis was restricted to these regions in order to serve as “background” frequencies for comparison to A′T methylated 4-mers, which are also mainly found downstream of TSSs. The 4-mers TATA and GATT are colored in red and blue, respectively, to facilitate comparison with methylated TA′TA and CA*TT in panel M. 
         FIGS. 11A-11D  show supplemental SMRT-seq data analyses. 
         FIG. 11A  shows the following: Top two panels depict PacBio coverage (horizontal axis) plotted against fractional methylation at each called 6 mA site (vertical axis). Bottom left panel is a histogram of fractional methylation of all 6 mA sites. Bottom right panel is a histogram of IPD ratios of all 6 mA sites. Mutant datasets show significantly lower fractional methylation and IPD ratios at 6 mA sites than wild-type data. 
         FIG. 11B  shows that wild-type SMRT-seq data are randomly subsampled 15 times, such that the resulting coverage is lower than ‘Mal mutant data. The difference in PacBio coverage between mutant and subsampled wild-type data is calculated for each chromosome, and is collectively represented as an olive boxplot (top panel). This set of calculations is repeated 15 times for each subsampled dataset, resulting in a series of 15 boxplots. The difference in PacBio coverage between mutant and fully sampled wild-type data is represented as a violet boxplot. Separately, the difference in total 6 mA marks per chromosome is calculated for respective datasets, and boxplots are shown in the bottom panel. Mutant datasets consistently yield lower numbers of called 6 mA marks than subsampled wild-type, despite the former having higher coverage than the latter. 
         FIG. 11C  shows the scatterplot of total number of 6 mA marks per chromosome in wild-type versus mutant data. PacBio cutoffs for calling 6 mA marks are varied as shown. A greater number of 6 mA marks per chromosome are consistently detected in wild-type than mutant data. 
         FIG. 11D  shows the boxplot of PacBio chromosome coverage in individual wild-type and mutant biological replicates (left panel). Only chromosomes with 100-150× PacBio coverage are shown. The total number of 6 mA marks in each of these chromosomes are plotted in the right panel. Wild-type replicates show consistently higher numbers of 6 mA marks per chromosome than mutant replicates. 
         FIGS. 12A-12H  show analysis of nucleosome organization and confirmation of ectopic DNA insertion in mta1 mutants. Description of analysis in panels A-G: Nucleosomes are grouped according to their “starting” 6 mA level, defined as the total number of 6 mA marks±200 bp from the nucleosome dyad in wild-type cells (WT). The dyad is assigned to be the peak position of MNase-seq reads. Similarly, linkers are grouped according to their “starting” methylation level, defined as the total number of 6 mA marks between two flanking nucleosome dyads (or between the 5′ chromosome end and the terminal nucleosome) in wild-type cells. Loci with high starting 6 mA have methylation greater than or equal to the 90th percentile of starting 6 mA levels, and show greater changes in methylation between mutant and wild-type cells ( FIG. 3D ). Those with low starting 6 mA are in the lowest 10th percentile. if 6 mA impacts nucleosome organization in vivo, then loci with high starting 6 mA should show a greater change in nucleosome organization. Possible effects are illustrated in panels A-C. Vertical green lines depict 6 mA marks, while blue and red peaks denote nucleosome occupancy. The plots shown in panels A-C illustrate the idealized result if 6 mA disfavors nucleosomes in vivo. Actual effects are shown in panels D-G. “Wild type” is abbreviated as WT. Analyses are restricted to the 5′ chromosome end. 
         FIG. 12A  shows that 6 mA loss may result in an increase in nucleosome fuzziness (highlighted with bold red double-sided arrow). The effect should be greater for nucleosomes with high starting 6 mA due to greater change in 6 mA between mutant and wild-type cells (“Change in nucleosome fuzziness” Box). Nucleosomes should, in turn, exhibit lower occupancy near the peak position, and higher occupancy in flanking regions (“Change in Nucleosome occupancy” Box; highlighted with red arrowheads and plotted ±73 bp from the dyad). Nucleosome fuzziness is calculated as the standard deviation of MNase-seq read locations ±73 bp from the dyad. 
         FIG. 12B  shows that 6 mA loss from nucleosome linker regions may result in a decrease in linker length (highlighted with bold red bracket). If so, the magnitude of decrease in linker length should be greater for linkers with high starting 6 mA (“Change in linker length” Box). 
         FIG. 12C  shows that 6 mA loss may result in an increase in occupancy directly over the methylated linker region (highlighted with bold red bracket). If so, the magnitude of increase in linker occupancy should be greater for regions with high starting 6 mA (“Change in linker occupancy” Box). Linker occupancy denotes the average MNase-seq coverage ±25 bp from the midpoint between flanking nucleosome dyads or chromosome end. As an example, for the +1/+2 nucleosome linker, occupancy is calculated ±25 bp from the midpoint of the +1 and +2 nucleosome dyad positions. Since nucleosome linker length in  Oxytricha  is ˜200 bp ( FIG. 12F , bottom panels), the genomic window used to calculate linker occupancy has minimal overlap with that for calculating nucleosome fuzziness and occupancy in panel A. 
         FIG. 12D  shows the impact of 6 mA loss on nucleosome fuzziness. For each nucleosome, the change in fuzziness between mutant and wild-type cells is calculated. Boxplots represent the distribution of changes in fuzziness scores. “MNase-seq” denotes sequencing of nucleosomal DNA obtained from  Oxytricha  chromatin in vivo, while “Control gDNA-seq” represents sequencing of MNase-digested, naked genomic DNA in vitro. Boxplot features are as described in  FIG. 1C . Distributions are compared using a Wilcoxon rank-sum test. N.S denotes “non-significant,” with p&gt;0.01. 
         FIG. 12E  shows the impact of 6 mA loss on nucleosome occupancy. For each nucleosome, the difference in nucleosome occupancy between mutant and wild-type cells is calculated at individual basepairs±73 bp around the nucleosome dyad. Data are averaged and depicted as line plots. The change in occupancy at the dyad is compared between nucleosomes with high and low starting 6 mA using a Wilcoxon rank-sum test. 
         FIG. 12F  shows the impact of 6 mA loss on linker length. Three types of linkers are analyzed: between the 5′ chromosome end and +1 nucleosome dyad, between the +1 and +2 nucleosome dyads, and between the +2 and +3 nucleosome dyads. For each linker, the difference in its length between mutant and wild-type cells is calculated. The resulting distribution of linker length differences is plotted as a histogram (top-most row of this panel). Distributions of linker length differences are compared using two-sample unequal variance t test. N.S. indicates “not significant,” with p&gt;0.01. Separately, the respective distributions of linker lengths in mutant and wild-type cells are plotted in the bottom two rows of this panel. The median linker length from each group is included as an inset. 
         FIG. 12G  shows the impact of 6 mA loss on linker occupancy. Linkers are binned as in panel F. For each linker, the difference in occupancy between mutant and wild-type cells is calculated. The resulting distribution of changes in linker occupancy is represented as a boxplot. Distributions are compared using two-sample unequal variance t test. N.S. indicates “not significant,” with p&gt;0.01. Boxplot features are as described in  FIG. 1C . 
         FIG. 12H  shows poly(A) +  RNaseq analysis of wild-type and mta1 mutants. “ATG” denotes start codon of MTA1 gene. A 62 bp ectopic DNA insertion results in a frameshift mutation in the MTA1 coding region. Three wild-type (WTI, WT2, wr3) and mutant (mta1′, mta12, mta13) biological replicates are analyzed. Short horizontal bars represent RNaseq reads, which are, −.75 nt in length and mapped to the reference sequence. For a read to be successfully mapped, it must have no more than 2 mismatches relative to the reference sequence. Unmapped reads are discarded. Blue and red bars denote RNaseq reads that map to native and ectopic regions, respectively. RNaseq reads overlapping the ectopic region are detected in mutant but not wild-type replicates. These reads span junctions between the ectopic and flanking coding regions, confirming the site of ectopic insertion. 
         FIGS. 13A-13I  show gel analysis of histone octamers and assembled chromatin. Description for panels A-D:  Xenopus  unmodified core histones were recombinantly expressed.  Oxytricha  histones were acid-extracted from vegetative nuclei.  Oxytricha  and  Xenopus  histones were subsequently refolded into octamers and purified through size exclusion chromatography. Description for panels E-I:  Xenopus  or  Oxytricha  histone octamers were assembled on DNA and subsequently digested with MNase to obtain ˜150 bp mononucleosome-sized fragments (labeled with red arrowheads). The resulting products were visualized by agarose gel electrophoresis. Mononucleosomal DNA was gel-excised and analyzed using Illumina sequencing or tiling qPCR analysis in  FIGS. 4A-4E, 6A — 6 E, and  14 A— 14 F. 
         FIG. 13A  shows reverse-phase HPLC purification of acid-extracted  Oxytricha  histones. Fractions 1-5 were individually collected and analyzed by Coomassie staining and western blotting. 
         FIG. 13B  shows SDS-PAGE analysis of purified  Oxytricha  histone fractions. 
         FIG. 13C  shows Western blot analysis of  Oxytricha  histone fractions 1-5. The fraction that is most enriched in each type of histone is colored in red. Arrowheads indicate likely histone bands. 
         FIG. 13D  shows SDS-PAGE analysis of purified  Oxytricha  and  Xenopus  histone octamers. 
         FIG. 13E  shows that chromatin was assembled on PCR-amplified  Oxytricha  mini-genome DNA, digested with MNase, and analyzed by agarose gel electrophoresis. 
         FIG. 13F  shows that chromatin was assembled on native  Oxytricha  genomic DNA, digested with MNase, and analyzed by agarose gel electrophoresis. 
         FIG. 13G  shows that chromatin was assembled with synthetic chromosome DNA, digested with MNase, and visualized by agarose gel electrophoresis. All assemblies with synthetic chromosomes were performed in the presence of an approximately 100-fold mass excess of buffer DNA relative to synthetic chromosome (see Example 1). This applies to panels G, H, and I. Representative assemblies with the unmethylated chromosome are shown. Methylated chromosome assemblies were separately performed in place of the unmethylated variant. 
         FIG. 13H  shows that chromatin was assembled on unmethylated synthetic chromosomes by salt dialysis and subsequently incubated with ACF and/or ATP. The resulting mixture was digested with MNase and visualized by agarose gel electrophoresis. Regularly spaced nucleosomes (labeled with red dots) are observed only when chromatin was incubated with both ACF and ATP. 
         FIG. 13I  shows chromatin assembled on unmethylated synthetic chromosomes using the NAP1 histone chaperone in the presence of ACF and/or ATP. The resulting mixture was digested with MNase and visualized by agarose gel electrophoresis. Nucleosomes are regularly spaced (labeled with red dots) in the presence of both ACF and ATP, although less apparent than in panel H. 
         FIGS. 14A-14F  show control MNase-Seq and tiling qPCR analysis. 
         FIG. 14A  is the same analysis as  FIG. 4C , showing that 6 mA quantitatively disfavors nucleosome occupancy in vitro but not in vivo. Here, the extent of MNase digestion was 40% of that in  FIG. 4C . P-values were calculated using a two-sample unequal variance t test. N.S denotes “non-significant,” with p&gt;0.05. 
         FIG. 14B  is the same analysis as  FIG. 6E , showing that the ACF complex restores nucleosome occupancy over methylated DNA in an ATP-dependent manner in vitro. Here, the extent of MNase digestion was 25% of that in  FIG. 6E . P-values were calculated using a two-sample unequal variance t test. N.S denotes “non-significant,” with p&gt;0.05. 
         FIG. 14C  is the same analysis as  FIG. 12D , showing that nucleosomes with high starting 6 mA show larger changes in fuzziness. Here, the extent of MNase digestion was 40% of that in  FIG. 12D . Distributions are compared using a Wilcoxon rank-sum test. N.S denotes “non-significant,” with p&gt;0.01. 
         FIG. 14D  is the same analysis as  FIG. 12E , showing that nucleosomes with high starting 6 mA exhibit characteristic changes in nucleosome occupancy at and around the nucleosome dyad. Here, the extent of MNase digestion was 40% of that in  FIG. 12E . The change in dyad occupancy is compared between nucleosomes with high and low starting 6 mA using a Wilcoxon rank-sum test. N.S denotes “non-significant,” with p&gt;0.01. 
         FIG. 14E  shows tiling qPCR analysis of nucleosome occupancy in spike-in and homogeneous synthetic chromosome preparations. The blunt, unmethylated synthetic chromosome (construct #1 in  FIG. 5B ) was used for chromatin assembly with (“Spike-in”) or without (“Homogeneous”) a 100-fold excess of buffer DNA. In the latter case, an equivalent mass of synthetic chromosome was added in place of buffer DNA to maintain the same DNA concentration for chromatin assembly. The tiling qPCR assay was performed as in  FIG. 6B . Shaded red bars depict the regions where 6 mA modulates nucleosome occupancy in separate methylated chromosomes analyzed in  FIGS. 6B and 6C . Note that methylated chromosomes were not used to generate qPCR data for this figure. Black arrowheads indicate no decrease in nucleosome occupancy in these regions when buffer DNA is used. Thus, the decrease in nucleosome occupancy in methylated chromosomes reported in  FIGS. 6A-6E  cannot be attributed to spike-in versus homogeneous addition of DNA for chromatin assembly. Error bars in all panels represent s.e.m. (n=3-4). 
         FIG. 14F  shows that chromatin was assembled on synthetic chromosomes using the NAP1 histone chaperone in the presence of ACF and/or ATP, instead of set dialysis. qPCR analysis was performed as in  FIG. 6B . Methylated chromosomes used in this experiment contain 6 mA in native sites. The addition of ACF and ATP results in a partial restoration of nucleosome occupancy over the methylated region. These results are similar to  FIG. 6D , where chromatin was assembled by sat dialysis instead of NAP1. 
         FIG. 15  shows that ciliate methyltransferase MTA1c mediates DNA N6-adenine methylation (6 mA) in vivo and 6 mA directly disfavors nucleosome occupancy in vitro. 
     
    
    
     DETAILED DESCRIPTION OF THE DISCLOSURE 
     DNA N6-adenine methylation (6 mA) has recently been described in diverse eukaryotes, spanning unicellular organisms to metazoa. In the present disclosure, it&#39;s reported a DNA 6 mA methyltransferase complex in ciliates, termed MTA1c. It consists of two MT-A70 proteins and two homeobox-like DNA-binding proteins and specifically methylates dsDNA. Disruption of the catalytic subunit, MTA1, in the ciliate  Oxytricha  leads to genome-wide loss of 6 mA and abolishment of the consensus ApT dimethylated motif. Mutants fail to complete the sexual cycle, which normally coincides with peak MTA1 expression. The present disclosure investigates the impact of 6 mA on nucleosome occupancy in vitro by reconstructing complete, full-length  Oxytricha  chromosomes harboring 6 mA in native or ectopic positions. It&#39;s shown that 6 mA directly disfavors nucleosomes in vitro in a local, quantitative manner, independent of DNA sequence. Furthermore, the chromatin remodeler ACF can overcome this effect. The present disclosure identifies a diverged DNA N6-adenine methyltransferase and defines the role of 6 mA in chromatin organization. 
     One embodiment of the present disclosure is a method of modifying a nucleic acid from a cell, the cell derived from a multicellular eukaryote. This method comprises the steps of: (a) obtaining the nucleic acid from the cell; and (b) contacting the nucleic acid with MTA1c or any components thereof under conditions effective to methylate the nucleic acid. 
     In some embodiments, the nucleic acid is RNA or DNA. In some embodiments, the eukaryotic cell is mammalian. In some embodiments, the multicellular eukaryote is a human. In some embodiments, the modification is a DNA N6-adenine methylation including one of more of the following motifs: dimethylated AT (5′-A*T-3′/3′-TA*-5′), dim ethylated TA (5′-TA*-3′/3′-A*T-5′), dim ethylated AA (5′-A*A*-3′/3′-TT-5′), methylated AT (5′-A*T-3′/3′-TA-5′), methylated AA (5′-A*A-3′/3′-TT-5′), methylated AC (5′-A*C-3′/3′-TG-5′), methylated AG (5′-A*G-3′/3′-TC-5′), methylated TA (5′-TA*-3′/3′-AT-5′), methylated AA (5′-AA*-3′/3′-TT-5′), methylated CA (5′-CA*-3′/3′-GT-5′), and methylated GA (5′-GA*-3′/3′-CT-5′). In certain embodiments, the MTA1 or an ortholog thereof comprises a mutation effective to abrogate dimethylation of the nucleic acid. Preferably, the mutation comprises loss of a C-terminal methyltransferase domain. In some embodiments, the MTA1c or any components thereof is obtained from ciliates, algae, or basal fungi. Preferably, the MTA1c or any components thereof is obtained from  Oxytricha  or  Tetrahymena.    
     As used herein, an “ortholog,” or orthologous gene, is a gene with a sequence that has a portion with similarity to a portion of the sequence of a known gene, but found in a different species than the known gene. An ortholog and the known gene originated by vertical descent from a single gene of a common ancestor. As used herein an ortholog encodes a protein that has a portion of at least about 50%, such as at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80% or at least about 80% of the total length of the sequence of the encoded protein that is similar to a portion of a length of at least about 50%, such as at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80% or at least about 80% of a known protein. The respective portion of the ortholog and the respective portion of the known protein to which it is similar may be a continuous sequence or be fragmented a number, for example, into 1 to about 3, including 2, individual regions within the sequence of the respective protein. For example, the 1 to about 3 regions are arranged in the same order in the amino acid sequence of the ortholog and the amino acid sequence of the known protein. Such a portion of an ortholog has an amino acid sequence that has at least about 40%, at least about 45%, such as at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75% or at least about 80% sequence identity to the amino acid sequence of the known protein encoded by a MTA1 gene. 
     As used herein, an asterisk “*” indicates the presence of a methylated base. For example, “A*” represents a methylated adenine. 
     The modified base, m6dA, has been discovered in a wide range of eukaryotes, including humans. m6dA levels are significantly reduced in gastric and liver cancer tissues, and disruption of m6dA promotes tumor formation (Xiao et al. 2018). As disclosed herein, MTA1 is a novel m6dA “writer”, paving the way for cost-effective methods to understand mechanisms of m6dA function in biomedically relevant models. 
     Accordingly, another embodiment of the present disclosure is a method of treating or ameliorating the effects of a disease characterized by an abnormal level of m6dA in a subject. This method comprises administering to the subject an amount of MTA1c or any components thereof effective to modulate m6dA levels in the subject. In some embodiments, the modulation comprises restoring m6dA levels to normal or near-normal ranges in the subject. 
     In some embodiments, the subject is a mammal that can be selected from the group consisting of humans, veterinary animals, and agricultural animals. Preferably, the subject is a human. 
     In some embodiments, the disease is a cancer, e.g., gastric cancer or liver cancer. In certain embodiments, the method further comprises administering to the subject one or more of anti-gastric cancer and anti-liver cancer drugs. Non-limiting examples of anti-liver cancer drugs include Nexavar™ (Sorafenib Tosylate) and Stivarga™ (Regorafenib). Non-limiting examples of anti-gastric cancer drugs include Cyramza™ (Ramucirumab), Doxorubicin Hydrochloride, 5-FU (Fluorouracil Injection), Fluorouracil Injection, Herceptin™ (Trastuzumab), Mitomycin C, Taxotere™ (Docetaxel), Trastuzumab, Afinitor™ (Everolimus), Somatuline Depot™ (Lanreotide Acetate), FU-LV, TPF, and XELIRI. 
     In some embodiments, the method furthering comprises co-administering to the subject an epigenetic agent that is selected from the group consisting of methylation inhibiting drugs, Bromodomain inhibitors, histone acetylase (HAT) inhibitors, protein methyltransferase inhibitors, histone methylation inhibitors, histone deacetlyase (HDAC) inhibitors, histone acetylases, histone deacetlyases, and combinations thereof. 
     Another embodiment of the present disclosure is a pharmaceutical composition comprising MTA1c or any components thereof that is effective to modulate m6dA levels in a subject in need thereof and a pharmaceutically acceptable carrier, diluent, adjuvant or vehicle. 
     Yet another embodiment of the present disclosure is a kit for treating or ameliorating the effects of a disease characterized by an abnormal level of m6dA in a subject, such as, e.g., cancer, comprising an effective amount of MTA1c or any components thereof, packaged together with instructions for its use. 
     Another embodiment of the present disclosure is a cell line obtained from a multicellular eukaryote comprising a nucleic acid encoding MTA1c or any components thereof and/or an MTA1c protein complex or any components thereof. As used herein, a “cell line” refers to all types of cell lines such as, e.g., immortalized cell lines and primary cell lines. In certain embodiments, the nucleic acid encoding MTA1c or any components thereof is operably linked to a recombinant expression vector. 
     Another embodiment of the present disclosure is a recombinant expression vector comprising a polynucleotide encoding MTA1c or any components thereof. 
     Still another embodiment of the present disclosure is a transgenic organism whose genome comprises a transgene comprising a nucleotide sequence encoding MTA1c or any components thereof. Non-limiting examples of possible organism include an archaea, a bacterium, a eukaryotic single-cell organism, algae, a plant, an animal, an invertebrate, a fly, a worm, a cnidarian, a vertebrate, a fish, a frog, a bird, a mammal, an ungulate, a rodent, a rat, a mouse, and a non-human primate. 
     The present disclosure also provides a method of identifying protein binding sites on DNA. This method comprises the steps of: (a) providing DNA; (b) contacting the DNA with MTA1c or any components thereof under conditions effective to methylate the DNA; (c) contacting the DNA with one or more proteins; (d) contacting the DNA with an enzyme effective to hydrolize the DNA in positions where no protein binding occurs; (e) removing the DNA bound protein; and (f) isolating and sequencing the DNA fragments. In certain embodiments, the one or more proteins in step (c) comprise histone octamers. 
     Another embodiment of the present disclosure is a method of mediating DNA N6-adenine methylation. This method comprises the steps of: (a) providing DNA; and (b) contacting the DNA with MTA1c or any components thereof under conditions effective to methylate the DNA. 
     Another embodiment of the present disclosure is a method of modulating nucleosome organization and/or transcription in a cell, comprising providing to the cell an agent that is effective to modulate the expression of MTA1c or any components thereof. 
     The present disclosure also provides a method of generating a synthetic chromosome. This method comprises the steps of: (a) generating chromosome segments containing terminal restriction sites, wherein the chromosome segments comprise one or more m6dA bases; (b) digesting the chromosome segments with a restriction enzyme; and (c) purifying and ligating the digested chromosome segments to form a synthetic chromosome. In some embodiments, the method further comprises enriching the synthetic chromosome. A synthetic chromosome made by the method above is also provided. 
     The following examples are provided to further illustrate certain aspects of the present disclosure. These examples are illustrative only and are not intended to limit the scope of the disclosure in any way. 
     EXAMPLES 
     Example 1 
     Materials and Methods 
       
     
       
         
           
               
            
               
                   
               
               
                 KEY RESOURCES TABLE 
               
            
           
           
               
               
               
            
               
                 REACIENT or RESOURCE 
                 SOURCE 
                 IDENTIFIER 
               
               
                   
               
            
           
           
               
            
               
                 Antibodies 
               
            
           
           
               
               
               
            
               
                 Anti-H2A 
                 Active Motif 
                 Cat #: 39111 
               
               
                 Anti-H2B 
                 Abcam 
                 Cat #: 1790 
               
               
                 Anti-H3 
                 Abcam 
                 Cat #: 1791 
               
               
                 Anti-H4 
                 Active Motif 
                 Cat #: 39269 
               
               
                 Anti-N6-methyladenosine 
                 Cedarlane 
                 Cat #: 202003(SY) 
               
               
                 antibody 
                 Labs/Synaptic 
               
               
                   
                 Systems 
               
               
                 Goat Anti-Rabbit IgG 
                 Bio-Rad 
                 1706515 
               
               
                 (H + L)-HRP Conjugate 
               
            
           
           
               
            
               
                 Bacterial and Virus Strains 
               
            
           
           
               
               
               
            
               
                 One Shot TOP10 chemically 
                 Thermo Fisher 
                 Cat #: C404006 
               
               
                 competent  E. coli   
               
               
                 BL21(DE3) pLysS 
                 Thermo Fisher 
                 Cat #: 70-236-4 
               
               
                 SHuffle T7 Express 
                 NEB 
                 Cat #: C3029J 
               
               
                 Competent  E. coli   
               
               
                 Lemo21 (DE3) Competent 
                 NEB 
                 Cat #: C2S28J 
               
               
                 
                   E. coli 
                 
               
            
           
           
               
            
               
                 Chemicals, Peptides, and Recombinant Proteins 
               
            
           
           
               
               
               
            
               
                 Micrococcal nuclease 
                 NEB 
                 Cat #: M0247S 
               
               
                 Q5 Site-Directed 
                 NEB 
                 Cat #: E0554S 
               
               
                 Mutagenesis Kit 
               
               
                 ProBlock Gold bacterial 
                 GoldBio 
                 Cat #: GB-330-5 
               
               
                 protease inhibitor 
               
               
                 cocktail 
               
               
                 Proteinase K 
                 Roche 
                 Cat #: 3113879001 
               
               
                 Phenol:Chloroform:IAA, 
                 Thermo Fisher 
                 Cat #: AM9732 
               
               
                 25:24:1 
               
               
                 TRIzol reagent 
                 Thermo Fisher 
                 Cat #: 15596026 
               
               
                 DNA Polymerase I, Large 
                 NEB 
                 Cat #: M0210S 
               
               
                 (Klenow) Fragment 
               
               
                 Klenow Fragment 
                 NEB 
                 Cat #: M0212S 
               
               
                 (3′ → 5′ exo-) 
               
               
                 Bsal 
                 NEB 
                 Cat #: R3535S 
               
               
                 EcoGII 
                 NEB 
                 Cat #: M0603S 
               
               
                 T4 DNA ligase 
                 NEB 
                 Cat #: M0202M 
               
               
                 Phusion DNA polymerase 
                 NEB 
                 Cat #: M0530L 
               
               
                 S-adenosyl-L-methionine 
                 NEB 
                 Cat #: B9003S 
               
               
                 Mouse NAP1 
                 This study 
                 N/A 
               
               
                 Drosophila ACF complex 
                 Active Motif 
                 Cat #: 31509 
               
               
                 Xenopus histones 
                 This study 
                 N/A 
               
               
                 Polyvinyl alcohol 
                 Sigma Aldrich 
                 Cat #: P8136 
               
               
                 Polyethylene glycol 8000 
                 Sigma Aldrich 
                 Cat #: P2139 
               
               
                 Adenosine 
                 Sigma Aldrich 
                 Cat #: A6559-25UMO 
               
               
                 5′-triphosphate (ATP) 
               
               
                 Creatine phosphate 
                 Sigma Aldrich 
                 Cat #: 10621714001 
               
               
                 Creatine kinase 
                 Sigma Aldrich 
                 Cat #: 10127566001 
               
               
                 Power SYBR Green PCR master 
                 Thermo Fisher 
                 Cat #: 4367659 
               
               
                 mix 
               
               
                 Gum Arabic 
                 Sigma Aldrich 
                 Cat #: G9752-1KG 
               
               
                 3H-labeled 
                 PerkinElmer 
                 Cat #: NET155V250UC 
               
               
                 S-adenosyl-L-methionine 
               
               
                 ([3H]SAM) 
               
               
                 Ultima Gold 
                 PerkinElmer 
                 Cat #: 6013326 
               
               
                 DNA degradase plus enzyme 
                 Zymo Research 
                 Cat #: E2020 
               
               
                   15 N 5 -dA nucleoside 
                 Cambridge 
                 Cat #: NLM-3895-25 
               
               
                   
                 Isotope 
               
               
                   
                 Laboratories 
               
               
                 D 3 -6mA 
                 Synthesized 
                 N/A 
               
               
                   
                 in this study 
               
            
           
           
               
            
               
                 Critical Commercial Assays 
               
            
           
           
               
               
               
            
               
                 QIAquick gel extraction kit 
                 QIAGEN 
                 Cat #: 28706 
               
               
                 NEBNext Poly(A) mRNA 
                 NEB 
                 Cat #: E7490S 
               
               
                 Magnetic Isolation 
               
               
                 Module 
               
               
                 ScriptSeq v2 RNA-Seq 
                 Illumina 
                 Cat #: SSV21124 
               
               
                 Library Prep Kit 
               
               
                 Nucleospin Tissue Kit 
                 Takara Bio 
                 Cat #: 740952.250 
               
               
                   
                 USA 
               
               
                 MinElute Reaction Cleanup 
                 QIAGEN 
                 Cat #: 28206 
               
               
                 Kit 
               
               
                 NEBNext Ultra II DNA 
                 NEB 
                 Cat #: E7645S 
               
               
                 Library Prep Kit 
               
               
                 Hi-Scribe T7 High Yield 
                 NEB 
                 Cat #: E2040S 
               
               
                 RNA Synthesis Kit 
               
               
                 Dynabeads Protein A 
                 Thermo Fisher 
                 Cat #: 10001D 
               
               
                 TOPO TA cloning kit 
                 Thermo Fisher 
                 Cat #: K457501 
               
            
           
           
               
            
               
                 Deposited Data 
               
            
           
           
               
               
               
            
               
                 Oxytricha trifallax 
                 This study 
                 SRA: SRX2335608 and 
               
               
                 SMRT-seq 
                   
                 SRX2335607 
               
               
                 Tetrahymena thermophila 
                 This study 
                 GEO: GSE94421 
               
               
                 SMRT-seq 
               
               
                 Oxytricha trifallax, all 
                 This study 
                 GEO: GSE94421 
               
               
                 Illumina data (RNA- 
               
               
                 seq, 6mA-IP-seq, MNase-seq, 
               
               
                 gDNA-seq) 
               
            
           
           
               
            
               
                 Experimental Models: Organisms/Strains 
               
            
           
           
               
               
               
            
               
                 Oxytricha trifallax cells, 
                 Lab collection 
                 N/A 
               
               
                 strain JRB310 
               
               
                 Oxytricha trifallax cells, 
                 Lab collection 
                 N/A 
               
               
                 strain JRB510 
               
               
                 Oxytricha trifallax cells, 
                 Lab collection 
                 N/A 
               
               
                 mtal mutant 
               
               
                 Tetrahymena thermophila 
                 Tetrahymena 
                 Cat #: SD00703 
               
               
                 cells, strain SB210 
                 stock center 
               
            
           
           
               
            
               
                 Oligonucleotides 
               
            
           
           
               
               
               
            
               
                 All are listed in Table S4 
                 IDT 
                 N/A 
               
            
           
           
               
            
               
                 Recombinant DNA 
               
            
           
           
               
               
               
            
               
                 pET-His-NAP1 (expression 
                 This study 
                 N/A 
               
               
                 vector for recombinant 
               
               
                 NAP1) 
               
               
                 pET-XenH2A (expression 
                 This study 
                 N/A 
               
               
                 vector for recombinant 
               
               
                 Xenopus histone H2A) 
               
               
                 pET-XenH2B (expression 
                 This study 
                 N/A 
               
               
                 vector for recombinant 
               
               
                 Xenopus histone H2B) 
               
               
                 pET-XenH3 (expression 
                 This study 
                 N/A 
               
               
                 vector for recombinant 
               
               
                 Xenopus histone H3) 
               
               
                 pET-XenH4 (expression 
                 This study 
                 N/A 
               
               
                 vector for recombinant 
               
               
                 Xenopus histone H4) 
               
               
                 pET-HisSUMO-MTA1 
                 This study 
                 N/A 
               
               
                 (expression vector for 
               
               
                 recombinant Tetrahymena 
               
               
                 MTA1) 
               
               
                 pET-HisSUMO-MTA7 
                 This study 
                 N/A 
               
               
                 (expression vector for 
               
               
                 recombinant Tetrahymena 
               
               
                 MTA7) 
               
               
                 pET-HisSUMO-p1 
                 This study 
                 N/A 
               
               
                 (expression vector for 
               
               
                 recombinant Tetrahymena p1) 
               
               
                 pET-HisSUMO-p2 
                 This study 
                 N/A 
               
               
                 (expression vector for 
               
               
                 recombinant Tetrahymena p2) 
               
               
                 pCR-TOPO- 
                 This study 
                 N/A 
               
               
                 syntheticChromosome (cloned 
               
               
                 synthetic chromosomes to 
               
               
                 verify accuracy of ligation 
               
               
                 of component DNA building 
               
               
                 blocks) 
               
            
           
           
               
            
               
                 Software and Algorithms 
               
            
           
           
               
               
               
            
               
                 Galaxy 
                 Galaxy 
                 https://usegalaxy.org/ 
               
               
                   
                 Community Hub 
               
               
                 Bowtie2 
                 Langmead and 
                 http://bowtie-bio.sourceforge.net/bowtie2/index.shtml 
               
               
                   
                 Salzberg, 2012 
               
               
                 TopHat2 
                 TopHat2 
                 https://ccb.jhu.edu/software/tophat/index.shtml 
               
               
                   
                 (Mortazavi et 
               
               
                   
                 al., 2008) 
               
               
                 Python 2.7.10 
                 Python Software 
                 https://www.python.org/download/releases/2.7/ 
               
               
                   
                 Foundation 
               
               
                 CAGEr 
                 Haberle et 
                 https://bioconductor.org/packages/release/bioc/html/CAGEr.html 
               
               
                   
                 al.. 2015 
               
               
                 SMRT Analysis 2.3.0 
                 Pacific 
                 https://www.pacb.com/documentation/smrt-analysis-software-installation-v2-3-0/ 
               
               
                   
                 Biosciences 
               
               
                 PSI-BLAST 
                 NCBI/NIH 
                 https://blast.ncbi.nlm.nih.gov/ 
               
               
                   
                   
                 Blast.cgi?CMD=Web&amp;PAGE-Proteins&amp;PROGRAM-blastp&amp;RUN_PSIBLAST=on 
               
               
                 CD-HIT 
                 Huang et al., 
                 http://weizhong-lab.ucsd.edu/cdhit-web-server/cgi-bin/index.cgi 
               
               
                   
                 2010 
               
               
                 MAFFT 
                 Katoh et al., 
                 https://mafft.cbrc.jp/alignment/software/ 
               
               
                   
                 2017; Kuraku 
               
               
                   
                 et al., 2013 
               
               
                 MrBayes/CIPRES Science 
                 Miller et al., 
                 https://www.phylo.org/ 
               
               
                 Gateway 
                 2010 
               
               
                 R (v3.2.5) 
                 The R Foundation 
                 https://www.r-project.org/ 
               
               
                 hmmscan 
                 Finn et al., 
                 https://www.ebi.ac.uk/Tools/hmmer/search/hmmscan 
               
               
                   
                 2015 
               
            
           
           
               
            
               
                 Other 
               
            
           
           
               
               
               
            
               
                 Agencourt Ampure XP beads 
                 Beckman Coulter 
                 Cat #: A63880 
               
               
                 Acid-extracted Oxytricha 
                 This study 
                 N/A 
               
               
                 histones 
               
               
                 Slide-A-Lyzer 3.5K MWCO 
                 Thermo Fisher 
                 Cat #: PI66110 
               
               
                 cassette 
               
               
                 Amersham Hybond-XL membrane 
                 GE Healthcare 
                 Cat #: RPN303S 
               
               
                 Amersham Hybond-N+ 
                 GE Healthcare 
                 Cat #: RPN119B 
               
               
                 membrane 
               
               
                 Volvic water 
                 Amazon 
                 https://www.amazon.com/Volvic-500m1-6-Pack/dp/B013PCK8M4/ 
               
               
                   
                   
                 ref=sr_1_1_a_it?_ie=UTF8&amp;qid=1538873999&amp;sr=8- 
               
               
                   
                   
                 1&amp;keyword_s=volvic&amp;dpID=418qEyu6yrUpreST=_SY300 
               
               
                   
                   
                 QL70 &amp;dpSrc=srch 
               
               
                   
               
            
           
         
       
       
       Oxytricha trifallax  
     
     Vegetative  Oxytricha trifallax  strain J RB310 was cultured at a density of 1.5×10 7  cells/L to 2.5×10 7  cells/L in Pringsheim media (0.11 mM Na 2 HPO 4 , 0.08 mM MgSO 4 , 0.85 mM Ca(NO 3 ) 2 , 0.35 mM KCl, pH 7.0) and fed daily with  Chlamydomonas reinhardtii . Cells were filtered through cheesecloth to remove debris and collected on a 10 pm Nitex mesh for subsequent experiments. 
     
       Tetrahymena thermophila  
     
     Stock cultures of vegetative  Tetrahymena thermophila  strain SB210 were maintained in Neff medium (0.25% w/v proteose peptone, 0.25% w/v yeast extract, 0.5% glucose, 33.3 pM FeCl 3 ). These cultures were inoculated into SSP medium (2% w/v proteose peptone, 0.1% w/v yeast extract, 0.2% w/v glucose, 33 pM FeCl 3 ) and grown to log-phase (˜3.5×10 5  cells/mL) through constant shaking at 125 rpm/30° C. 
     In Vivo MNase-Seq 
     3×10 5  vegetative  Oxytricha  cells were fixed in 1% w/v formaldehyde for 10 min at room temperature with gentle shaking, and then quenched with 125 mM glycine. Cells were lysed by dounce homogenization in lysis buffer (20 mM Tris pH 6.8, 3% w/v sucrose, 0.2% v/v Triton X-100, 0.01% w/v spermidine trihydrochloride) and centrifuged in a 10%-40% discontinuous sucrose gradient (Lauth et al., 1976) to purify macronuclei. The resulting macronuclear preparation was pelleted by centrifugation at 4000×g, washed in 50 ml TMS buffer (10 mM Tris pH 7.5, 10 mM MgCl 2 , 3 mM CaCl 2 ), 0.25M sucrose), resuspended in a final volume of 300 μL, and equilibriated at 37° C. for 5 min. Chromatin was then digested with MNase (New England Biolabs) at a final concentration of 15.7 Kunitz Units/μL at 37° C. for 1 min 15 s, 3 min, 5 min, 7 min 30 sec, 10 min 30 s, and 15 min respectively. Reactions were stopped by adding ½ volume of PK buffer (300 mM NaCl, 30 mM Tris pH 8, 75 mM EDTA pH 8, 1.5% w/v SDS, 0.5 mg/mL Proteinase K). Each sample was incubated at 65° C. overnight to reverse crosslinks and deproteinate samples. Subsequently, nucleosomal DNA was purified through phenol:chloroform:isoamyl alcohol extraction and ethanol precipitation. Each sample was loaded on a 2% agarose-TAE gel to check the extent of MNase digestion. The sample exhibiting −80% mononucleosomal species was selected for MNase-seq analysis, in accordance with previous guidelines (Zhang and Pugh, 2011). Mononucleosome-sized DNA was gel-purified using a QIAquick gel extraction kit (QIAGEN). Illumina libraries were prepared using an NEBNext Ultra II DNA Library Prep Kit (New England Biolabs) and subjected to paired-end sequencing on an Illumina HiSeq 2500 according to manufacturer&#39;s instructions. All vecietative  Tetrahymena  MNase-sea data were obtained from (Beh et al., 2015). 
     Poly(A) +  RNA-Seq and TSS Sequencing 
       Oxytricha  cells were lysed in TRIzol reagent (Thermo Fisher Scientific) for total RNA isolation according to manufacturer&#39;s instructions. Poly(A) +  RNA was then purified using the NEBNext Poly(A) mRNA Magnetic Isolation Module (New England Biolabs).  Oxytricha  poly(A) +  RNA was prepared for RNA-seq using the ScriptSeq v2 RNA-Seq Library Preparation Kit (Illumina).  Tetrahymena  poly(A) +  RNA-seq data was obtained from (Xiong et al., 2012). The 5′ ends of capped RNAs were enriched from vegetative  Oxytricha  total RNA using the RAMPAGE protocol (Batut et al., 2013), and used for library preparation, Illumina sequencing and subsequent transcription start site determination (ie. “TSS-seq”). These data were used to plot the distribution of  Oxytricha  TSS positions in  FIG. 1A . TSS positions used for analysis outside of  FIG. 1A  were obtained from (Swart et al., 2013) and (Beh et al., 2015). For RNaseq analysis of genes grouped according to “starting” methylation level level: total 6 mA was counted between 100 bp upstream to 250 bp downstream of the TSS. Genes with high starting methylation have total 6 mA in the 90th percentile and higher. Genes with low starting methylation have total 6 mA at or below the 10th percentile. 
     Immunoprecipitation and Illumina Sequencing of Methylated DNA (6 mA IP-Seq) 
     Genomic DNA was isolated from vegetative  Oxytricha  cells using the Nucleospin Tissue Kit (Takara Bio USA, Inc.). DNA was sheared into 150 bp fragments using a Covaris LE220 ultra-sonicator (Covaris). Samples were gel-purified on a 2% agarose-TAE gel, blunted with DNA polymerase I (New England Biolabs), and purified using MinElute spin columns (QIAGEN). The fragmented DNA was dA-tailed using Klenow Fragment (3′-&gt;5′ exo-) (New England Biolabs) and ligated to Illumina adaptors following manufacturer&#39;s instructions. Subsequently, 2.2 μg of adaptor-ligated DNA containing 6 mA was immunoprecipitated using an anti-N6-methyladenosine antibody (Cedarlane Labs) conjugated to Dynabeads Protein A (Invitrogen). The anti-6 mA antibody is commonly used for RNA applications, but has also been demonstrated to recognize 6 mA in DNA (Fioravanti et al., 2013; Xiao and Moore, 2011). The immunoprecipitated and input libraries were treated with proteinase K, extracted with phenol:chloroform, and ethanol precipitated. Finally, they were PCR-amplified using Phusion Hot Start polymerase (New England Biolabs) and used for Illumina sequencing. 
     Sample Preparation for SMRT-Seq 
     Vegetative  Oxytricha  macronuclei were isolated as described in the subheading “in vivo MNase-seq” of this study. Vegetative  Tetrahymena  macronuclei were isolated by differential centrifugation (Beh et al., 2015).  Oxytricha  and  Tetrahymena  cells were not fixed prior to nuclear isolation. Genomic DNA was isolated from  Oxytricha  and  Tetrahymena  macronuclei using the Nucleospin Tissue Kit (Macherey-Nagel). Alternatively, whole  Oxytricha  cells instead of macronuclei were used. SMRT-seq according to manufacturer&#39;s instructions, using P5-C3 and P6-C4 chemistry, as in (Chen et al., 2014).  Oxytricha  and  Tetrahymena  macronuclear DNA were used for SMRT-seq in  FIGS. 1A-1E and 9A-9F , while  Oxytricha  whole cell DNA was used for all other Figures. Since almost all DNA in  Oxytricha  cells is derived from the macronucleus (Prescott, 1994), similar results are expected between the use of purified macronuclei or whole cells. 
     Illumina Data Processing 
     Reads from all biological replicates were merged before downstream processing. All Illumina sequencing data were quality trimmed (minimum quality score=20) and length-filtered (minimum read length=40nt) using Galaxy (Blankenberg et al., 2010; Giardine et al., 2005; Goecks et al., 2010). MNase-seq and 6 mA IP-seq reads were mapped to complete chromosomes in the  Oxytricha trifallax  JRB310 (August 2013 build) or  Tetrahymena thermophila  SB210 macronuclear reference genomes (June 2014 build) using Bowtie2 (Langmead and Salzberg, 2012) with default settings, while poly(A). RNA-seq and TSS-seq reads were mapped using TopHat2 (Mortazavi et al., 2008) with August 2013  Oxytricha  gene models or June 2014  Tetrahymena  gene models, with default settings. 
     MNase-seq datasets were generated by paired-end sequencing. Within each MNase-seq dataset, the read pair length of highest frequency was identified. All read pairs with length±25 bp from this maximum were used for downstream analysis. On the other hand, 6 mA IP-seq datasets were generated by single-read sequencing. 6 mA IP-seq single-end reads were extended to the mean fragment size, computed using cross-correlation analysis (Kharchenko et al., 2008). The per-basepair coverage of  Oxytricha  MNase-seq read pair centers and extended 6 mA IP-seq reads were respectively computed across the genome. Subsequently, the per-basepair coverage values were normalized by the average coverage within each chromosome to account for differences in DNA copy number (and hence, read depth) between  Oxytricha  chromosomes (Swart et al., 2013). The per-basepair coverage values were then smoothed using a Gaussian filter of standard deviation=15. This smoothed data is denoted as “normalized coverage” or “nucleosome occupancy.”  Tetrahymena  MNase-seq data were processed similarly to  Oxytricha , except that DNA copy number normalization was omitted as  Tetrahymena  chromosomes have uniform copy number (Eisen et al., 2006). 
     For the MNase-seq analysis in  FIGS. 4C, 6E, 14A, and 14B , nucleosome occupancy and 6 mA IP-seq coverage were calculated within overlapping 51 bp windows across the 98 assayed chromosomes. Windows were binned according to the number of 6 mA residues within. The in vitro MNase-seq coverage from chromatinized native gDNA (“+” 6 mA) was divided by the corresponding coverage from chromatinized mini-genome DNA (“−” 6 mA) to obtain the fold change in nucleosome occupancy in each window. Alternatively, a subtraction was performed on these datasets to obtain the difference in nucleosome occupancy in vitro. Identical DNA sequences were compared for each calculation. These data are labeled as (“+” histones) in  FIGS. 4C and 14A . Naked native gDNA and mini-genome DNA were also MNase-digested, sequenced and analyzed in the same manner to control for Mnase sequence preferences (“−” histones). Nucleosome occupancy in vivo corresponds to normalized MNase-seq coverage from wild type and mta1 mutant cells. 
     Nucleosome positions were iteratively called as local maxima in normalized MNase-seq coverage, as previously described (Beh et al., 2015). “Consensus”+1, +2, +3 nucleosome positions downstream of the TSS were inferred from aggregate MNase-seq profiles across the genome ( FIG. 1A  for  Oxytricha  and  FIG. 9A  for  Tetrahymena ). Each gene was classified as having a +1, +2, +3 and/or +4 nucleosome if there is a called nucleosome dyad within 75 bp of the consensus nucleosome position. 
     RNA-seq and TSS-seq read coverage were calculated without normalization by DNA copy number since there is no correlation between  Oxytricha  DNA and transcript levels (Swart et al., 2013). 
       Oxytricha  TSSs were called from TSS-seq data using CAGEr (Haberle et al., 2015); with clusterCTSS parameters (threshold=1.6, thresholdlsTpm=TRUE, nrPassThreshold=1, method=“paraclu,” removeSingletons=TRUE, keepSingletonsAbove=5). Only TSSs with tags per million counts&gt;0.1 were used for downstream analysis.  Tetrahymena  TSSs were obtained from (Beh et al., 2015). 
     SMRT-Seq Data Processing 
     We processed SMRT-seq data with SMRTPipe v1.87.139483 in the SMRT Analysis 2.3.0 environment using, in order, the P Fetch, P Filter (with minLength=50, minSubreadLength=50, readScore=0.75, and artifact=−1000), P FilterReports, P Mapping (with gff2Bed=True, pulsemetrics=DeletionQV, IPD, InsertionQV, PulseWidth, QualityValue, MergeQV, SubstitutionQV, DeletionTag, and load PulseOpts=byread), P_MappingReports, P_GenomicConsensus (with algorithm=quiver, outputConsensus=True, and enableMapQVFilter=True), P_ConsensusReports, and P Mod ificationDetection (with identifyModifcations=True, enableMapQVFilter=False, and mapQvThreshold=10) modules. All other parameters were set to the default. The  Oxytricha  August 2013 reference genome build was used for mapping  Oxytricha  SMRT-seq reads, with Contig10040.0.1, Contig1527.0.1, Contig4330.0.1, and Contig54.0.1 removed, as they are perfect duplicates of other Contigs in the assembly.  Tetrahymena  SMRT-seq reads were mapped to the June 2014 reference genome build. Only chromosomes with high SMRT-seq coverage (&gt;=80× for  Oxytricha; &gt;= 100× for  Tetrahymena ) were used for all 6 mA-related analyses. 
     Chromosome Synthesis 
     Synthetic Contig1781.0 chromosomes were constructed from “building blocks” of native chromosome sequence ( FIGS. 5B and 5C ). The dark blue building block in  FIG. 5B  was prepared by annealing synthetic oligonucleotides, while all other building blocks were generated by PCR-amplification from genomic DNA using Phusion DNA polymerase (New England Biolabs). All oligonucleotides used for annealing and PCR amplification are listed in Table 2. The PCR-amplified building blocks contain terminal restriction sites for BsaI (New England Biolabs), a type IIS restriction enzyme that cuts distal from these sites. BsaI cleaves within the native DNA sequence, generating custom 4nt 5′ overhangs and releasing the non-native BsaI restriction site as small fragments that are subsequently purified away. The BsaI-generated overhangs are complementary only between adjacent building blocks, conferring specificity in ligation and minimizing undesired by-products. After BsaI digestion, PCR building blocks were purified by phenol:chloroform extraction and ethanol precipitation. Building blocks were then sequentially ligated to each other using T4 DNA ligase (New England Biolabs) and purified by phenol:chloroform extraction and ethanol precipitation. Size selection after each ligation step was performed using polyethylene glycol (PEG) precipitation or Ampure XP beads (Beckman Coulter) to enrich for the large ligated product over its smaller constituents. The size of individual building blocks and their corresponding order of ligation were designed to maximize differences in size between ligated products and individual building blocks. This increases the efficiency in size selection of products over reactants. Chromosomes 1 and 6 in  FIG. 5B  was generated by full length PCR from genomic DNA. To prepare chromosomes 2-4 in  FIG. 5B , the red, dark blue, and purple blocks were first ligated in a 3-piece reaction and purified from the individual components. This product was subsequently ligated with the turquoise building block to obtain the full length chromosome. To prepare chromosomes 5 in  FIG. 5B , the red, orange, and emerald building blocks were ligated in a 3-piece reaction and subsequently purified. All chromosomes were subjected to Sanger sequencing to verify ligation junctions. 6 mA was installed in synthetic chromosomes using annealed oligonucleotides, or by incubation of DNA building blocks with EcoGII methyltransferase (New England Biolabs). 
     Verification of Synthetic Chromosome Sequences 
     All chromosomes were dA-tailed using Klenow Fragment (3′-&gt;5′ exo-) (New England Biolabs), cloned using a TOPO TA cloning kit (Thermo Fisher) or StrataClone PCR Cloning Kit (Agilent Technologies), transformed into One Shot TOP10 chemically competent  E. coli , and sequenced using flanking T7, T3, M13F, or M13R primers. 
     Preparation of  Oxytricha  Histones 
     Vegetative  Oxytricha trifallax  strain JRB310 was cultured as described in the subheading: “Experimental model and subject details” of this study. Cells were starved for 14 hr and subsequently harvested for macronuclear isolation as described in the subheading: “in vivo MNase-seq” of this study. However, formaldehyde fixation was omitted. Purified nuclei were pelleted by centrifugation at 4000×g, resuspended in 0.421 mL 0.4N H 2 SO 4  per 10 6  input cells, and nutated for 3 hr at 4° C. to extract histones. Subsequently, the acid-extracted mixture was centrifuged at 21,000× a for 15 min to remove debris. Proteins were precipitated from the cleared supernatant using trichloroacetic acid (TCA), washed with cold acetone, then dried and resuspended in 2.5% v/v acetic acid. Individual core histone fractions were purified from crude acid-extracts using semi-preparative RP-HPLC (Vydac C18, 12 micron, 10 mM×250 mm) with 40%-65% HPLC solvent B over 50 min ( FIG. 13A ). The identity of each purified histone fraction was verified by western analysis ( FIG. 13C ) using antibodies: anti-H2A (Active Motif #39111), anti-H2B (Abcam #ab1790), anti-H3 (Abcam #ab1791), anti-H4 (Active Motif #39269). 
     Preparation of Recombinant  Xenopus  Histones 
     All RP-HPLC analyses were performed using 0.1% TFA in water (HPLC solvent A), and 90% acetonitrile, 0.1% TFA in water (HPLC solvent B) as the mobile phases. Wild-type  Xenopus  H4, H3 C110A, H2B and H2A proteins were expressed in BL21(DE3) pLysS  E. coli  and purified from inclusion bodies through ion exchange chromatography (Debelouchina et al., 2017). Purified histones were characterized by ESI-MS using a MicrOTOF-Q II ESI-Qq-TOF mass spectrometer (Bruker Daltonics). H4: calculated 11,236 Da, observed 11,236.1 Da; H3 C110A: calculated 15,239 Da, observed 15,238.7 Da; H2A: calculated 13,950 Da, observed 13,949.8 Da; H2B: calculated 13,817 Da, observed 13,816.8 Da. 
     Preparation of Histone Octamers 
       Oxytricha  and  Xenopus  histone octamers were respectively refolded from core histones using established protocols (Beh et al., 2015; Debelouchina et al., 2017). Briefly, lyophilized histone proteins ( Xenopus  modified or wild-type;  Oxytricha  acid-extracted) were combined in equimolar amounts in 6 M guanidine hydrochloride, 20 mM Tris pH 7.5 and the final concentration was adjusted to 1 mg/mL. The solution was dialyzed against 2M NaCl, 10 mM Tris, 1 mM EDTA, and the octamers were purified from tetramer and dimer species using size-exclusion chromatography on a Superdex 200 10/300 column (GE Healthcare Life Sciences). The purity of each fraction was analyzed by SDS-PAGE. Pure fractions were combined, concentrated and stored in 50% v/v glycerol at −20° C. 
     Preparation of Mini-Genome DNA 
     98 full-length chromosomes were individually amplified from  Oxytricha trifallax  strain JRB310 genomic DNA using Phusion DNA polymerase (New England Biolabs). Primer pairs are listed in Table 2. Amplified chromosomes were separately purified using a MinElute PCR purification kit (QIAGEN), and then mixed in equimolar ratios to obtain “mini-genome” DNA. The sample was concentrated by ethanol precipitation and adjusted to a final concentration of ˜1.6 mg/mL. 
     Preparation of Native Genomic DNA for Chromatin Assembly Starry 
     Macronuclei were isolated from vegetative  Oxytricha trifallax  strain JRB310 as described in the subheading “in vivo MNase-seq” of this study. However, cells were not fixed prior to nuclear isolation. Genomic DNA was purified using the Nucleospin Tissue kit (Macherey-Nagel). Approximately 200 μg of genomic DNA was loaded on a 15%-40% linear sucrose gradient and centrifuged in a SW 40 Ti rotor (Beckman Coulter) at 160,070×g for 22.5 hr at 20° C. Sucrose solutions were in 1M NaCl, 20 mM Tris pH 7.5, 5 mM EDTA. Individual fractions from the sucrose gradient were analyzed on 0.9% agarose-TAE gels. Fractions containing high molecular weight DNA that migrated at the mobility limit were discarded as such DNA species were found to interfere with downstream chromatin assembly. All other fractions were pooled, ethanol precipitated, and adjusted to 0.5 mg/mL DNA. 
     Chromatin Assembly and Preparation of Mononucleosomal DNA 
     Chromatin assemblies were prepared by salt gradient dialysis as previously described (Beh et al., 2015; Luger et al., 1999), or using mouse NAP1 histone chaperone and  Drosophila  ACF chromatin remodeler as previously described (An and Roeder, 2004; Fyodorov and Kadonaga, 2003). Details of each chromatin assembly procedure are listed below. To reduce sample requirements while maintaining adequate DNA concentrations for chromatin assembly, synthetic chromosomes were first mixed with a hundred-fold excess of “buffer” DNA (PCR-amplified  Oxytricha  Contig17535.0). We verified that nucleosome occupancy in the methylated region (qPCR primer pairs 6 and 7) of the synthetic chromosome is unaffected by the presence of buffer DNA ( FIG. 14E ). Native and mini-genome DNA were not mixed with buffer DNA prior to chromatin assembly. 
     For chromatin assembly through salt dialysis: histone octamers and (synthetic chromosome+buffer) DNA were mixed in a 0.8:1 mass ratio, while histone octamers and (native or mini-genome) DNA were mixed in a 1.3:1 mass ratio, each in a 50 μL total volume. Samples were first dialyzed into start buffer (10 mM Tris pH 7.5, 1.4M KCl, 0.1 mM EDTA pH 7.5, 1 mM DTT) for 1 hr at 4° C. Then, 350 mL end buffer (10 mM Tris pH 7.5, 10 mM KCl, 0.1 mM EDTA, 1 mM DTT) was added at a rate of 1mUmin with stirring. The assembled chromatin was dialyzed overnight at 4° C. into 200 mL end buffer, followed by a final round of dialysis in fresh 200 mL end buffer for 1 hr at 4° C. The assembled chromatin was then adjusted to 50 mM Tris pH 7.9, 5 mM CaCl 2 ) and digested with MNase (New England Biolabs) to mainly mononucleosomal DNA as previously described (Beh et al., 2015). 
     For chromatin assembly using mouse NAP1 and  Drosophila  ACF: NAP1 was recombinantly expressed and purified as described in (An and Roeder, 2004). ACF was purchased from Active Motif. 0.49 μM NAP1 and 58 nM histone octamer were first mixed in a 302p1 reaction volume containing 62 mM KCl, 1.2% w/v polyvinyl alcohol (Sigma Aldrich), 1.2% w/v polyethylene glycol 8000 (Sigma Aldrich), 25 mM HEPES-KOH pH 7.5, 0.1 mM EDTA-KOH, 10% v/v glycerol, and 0.01% v/v NP-40. The NAP1-histone mix was incubated on ice for 30 min. Meanwhile, “AM” mix was prepared, consisting of 20 mM ATP (Sigma Aldrich), 200 mM creatine phosphate (Sigma Aldrich). 33.3 mM MgCl 2 , 33.3 μg/μl creatine kinase (Sigma Aldrich) in a 56u1 reaction volume. After the 30 min incubation. 5.29 μl of 1.7 μM ACF complex (Active Motif) and the “AM” mix were sequentially added to the NAP1-histone mix. Then, 10.63 μl of native or mini-genome DNA (2.66 μg) was added, resulting in a 374 μl reaction volume. The final mixture was incubated at 27° C. for 2.5 hr to allow for chromatin assembly. Subsequently, CaCl 2  was added to a final concentration of 5 mM, and the chromatin was digested with MNase (New England Biolabs) to mainly mononucleosomal DNA as previously described (Beh et al., 2015). 
     Mononucleosome-sized DNA from MNase-digested chromatin was gel-purified and used for tiling qPCR on a Viia 7 Real-Time PCR System with Power SYBR Green PCR master mix (Thermo Fisher), or in vitro MNase-seq on an Illumina HiSeq 2500, according to the manufacturer&#39;s instructions. qPCR primer sequences are listed in Table 2. 
     Tiling qPCR Analysis of Nucleosome Occupancy 
     qPCR data were analyzed using the ΔΔCt method (Livak and Schmittgen, 2001). At each locus along the synthetic chromosome, ΔCt=(Ct at locus of interest)−(Ct at qPCR primer pair 22, far from the methylated region). See  FIG. 6B  for location of qPCR primer pair 22. Separate ΔCt values were calculated from mononucleosomal DNA and the corresponding naked, undigested synthetic chromosome. The ΔΔCt value was calculated from this pair of ΔCt values. This controls for potential variation in PCR amplification efficiency, especially over methylated regions. The fold change in mononucleosomal DNA relative to naked chromosomal DNA at a particular locus is calculated as 2 −ΔΔCt , and denotes ‘nucleosome occupancy’ for all presented qPCR data. 
     ACF Spacing Assay 
     ATP-dependent nucleosome spacing was performed in accordance with a previous study (Lieleg et al., 2015). Chromatin was assembled by salt gradient dialysis as described above, and then adjusted to 20 mM HEPES-KOH pH 7.5, 80 mM KCl, 0.5 mM EGTA, 12% v/v glycerol, 10 mM (NH 4 ) 2 SO 4 , 2.5 mM DTT. Samples were then incubated for 2.5 hr at 27° C. with 3 mM ATP, 30 mM creatine phosphate, 4 mM MgCl 2 , 5 ng/0 creatine kinase, and 11 ng/μL ACF complex (Active Motif). Remodeled chromatin was then adjusted to 5 mM CaCl 2 ) and subjected to MNase digestion, mononucleosomal DNA purification, and qPCR analysis as described above. 
     Phylogenetic Analysis 
     The MTA1 amino acid sequence (UniProt ID: J9IF92 9SPIT) was queried against the NCBI nr database using PSI-BLAST (Altschul et al., 1997; Schaffer et al., 2001) (maximum e-value=1e −4 ; enable short queries and filtering of low complexity regions). Retrieved hits were collapsed using CD-HIT (Huang et al., 2010) with minimum sequence identity=0.97 to remove redundant sequences. The resulting sequences were added to existing MT-A70 alignments from (Greer et al., 2015) using MAFFT (-add) (Katoh et al., 2017; Kuraku et al., 2013). Gaps and duplicate sequences were removed from the merged alignment. Only sequences corresponding to the taxa in  FIG. 2B  were retained. The alignment was then used for phylogenetic tree construction using MrBayes in the CIPRES Science Gateway (Miller et al., 2010) with 5×10 6  generations. Protein sequences used for MrBayes analysis are given in Table 1. 
     The above procedure was also used for constructing phylogenetic trees from p1 (UniProt ID: Q22VV9 TETTS) and p2 (UniProt ID: I7M8B9 TETTS). However, protein sequences were aligned using MAFFT without adding to an existing alignment. 
     Preparation of Nuclear Extracts with DNA Methyltransferase Activity 
     Vegetative  Tetrahymena  cells were grown in SSP medium to log-phase (˜3.5×10 6  cells/mL) and collected by centrifugation at 2,300×g for 5 min in an SLA-3000 rotor. The supernatant was discarded, and cells were resuspended in medium B (10 mM Tris pH 6.75, 2 mM MgCl 2 , 0.1M sucrose, 0.05% w/v spermidine trihydrochloride, 4% w/v gum Arabic, 0.63% w/v 1-octanol, and 1 mM PMSF). Gum arabic (Sigma Aldrich) is prepared as a 20% w/v stock and centrifuged at 7,000×g for 30 min to remove undissolved clumps. For each volume of cell culture, one-third volume of medium B was added to the  Tetrahymena  cell pellet. Cells were resuspended and homogenized in a chilled Waring Blender (Waring PBB212) at high speed for 40 s. The resulting lysate was subsequently centrifuged at 2,750×g for 5 min in an SLA-3000 rotor to pellet macronuclei. The nuclear pellet was washed twice with medium B and then five times in MM medium (10 mM Tris-HCl pH 7.8, 0.25M sucrose, 15 mM MgCl 2 , 0.1% w/v spermidine trihydrochloride, 1 mM DTT, 1 mM PMSF). Macronuclei were pelleted between wash steps by centrifuging at 2,500×g for 5 min in an SLA-3000 rotor. Finally, the total number of washed macronuclei was counted with a hemocytometer using a Zeiss ID03 microscope. Nuclear proteins were extracted by vigorously resuspending the pellet in M M salt buffer (10 mM Tris-HCl pH 7.8, 0.25M sucrose, 15 mM MgCl2, 350 mM NaCl, 0.1% w/v spermidine trihydrochloride, 1 mM DTT, 1 mM PMSF). 1 mL M M salt buffer was added per 2.33×108 macronuclei. The viscous mixture was nutated for 45 min at 4° C., and then cleared at 175,000×g for 30 min at 4° C. in a SW 41 Ti rotor. Following this, the supernatant was dialyzed in a Slide-A-Lyzer 3.5K MWCO cassette (Thermo Fisher) overnight at 4° C. against two changes of MM minus medium (10 mM Tris-HCl pH 7.8, 15 mM MgCl 2 , 1 mM DTT, 0.5 mM PMSF). The dialysate was then centrifuged at 7,197×g for 1 hr at 4″C to remove precipitates, and dialyzed overnight in a Slide-A-Lyzer 3.5K MWCO cassette (Thermo Fisher) at 4° C. against two changes of MN3 buffer (30 mM Tris-HCl pH 7.8, 1 mM EDTA, 15 mM NaCl, 20% v/v glycerol, 1 mM DTT, 0.5 mM PMSF). The final dialysate was cleared by centrifugation at 7,197 g for 1.5 hr at 4° C., flash frozen, and stored at −80° C. This nuclear extract was used for all subsequent biochemical fractionation and 6 mA methylation assays. 
     Partial Purification of MTA1c from Nuclear Extracts 
       Tetrahymena  nuclear extracts were passed through a HiTrap O HP column (GE Healthcare) and eluted using a linear aradient of 15 mM to 650 mM NaCl in 30 mM Tris-HCl pH 7.8, 1 mM EDTA, 20% v/v glycerol, 1 mM DTT, 0.5 mM PMSF, over 30 column volumes. Each fraction was assayed for DNA methyltransferase activity using radiolabeled SAM as described in the next section. The DNA methyltransferase activity eluted in two peaks, at ˜60 mM and ˜365 mM NaCl, termed the “low salt sample” and “high salt sample.” Fractions corresponding to each peak were pooled and passed through a HiTrap Heparin HP column (GE Healthcare). Bound proteins were eluted using a linear gradient of 60 mM to 1M NaCl (for the low salt sample) or 350 mM to 1M NaCl (for the high salt sample) over 30 column volumes. Fractions with DNA methyltransferase activity were respectively pooled and dialyzed into 10 mM sodium phosphate pH 6.8, 100 mM NaCl, 10% v/v glycerol, 0.3 mM CaCl 2 ), 0.5 mM DTT (for the low salt sample); or 30 mM Tris-HCl pH 7.8, 1 mM EDTA, 200 mM NaCl, 10% v/v glycerol, 1 mM DTT, 0.2 mM PMSF (for the high salt sample). The dialyzed low salt sample was passed through a Nuvia cPrime column (Bio-Rad) and eluted using a linear gradient of 100 mM to 1M NaCl in 50 mM sodium phosphate pH 6.8, 10% v/v glycerol, 0.5 mM DTT. Separately, the dialyzed high salt sample was fractionated using a Superdex 200 10/300 GL column (GE Healthcare) in 30 mM Tris-HCl pH 7.8, 1 mM EDTA, 200 mM NaCl, 10% v/v glycerol, 1 mM DTT. Fractions from the Nuvia cPrime and Superdex 200 columns were dialyzed into 30 mM Tris-HCl pH 7.8, 1 mM EDTA, 15 mM NaCl, 20% v/v glycerol, 1 mM DTT, 0.5 mM PMSF and assayed for DNA methyltransferase activity. Those with qualitatively low, medium, and high activity were subjected to mass spectrometry to identify candidate methyltransferase proteins ( FIG. 2D ; Table 6). This experiment identified four proteins that co-purify with DNA methyltransferase activity—MTA1, MTA9, p1, and p2—and are collectively termed as “MTA1c” in the present disclosure. All four proteins are necessary for 6 mA methylation in vitro. 
     Recombinant Expression of MTA1, MTA9, p1, and p2 Proteins 
     Full length MTA1, MTA9, p1, and p2 open reading frames were codon-optimized for bacterial expression and cloned into a pET-His6-SUMO vector using ligation independent cloning. Protein sequences are listed in Table 3. The vector was a gift from Scott Gradia (Addgene plasmid #29659; http://addgene.org/29659; RRID: Addgene 29659). Mutations in the MTA1 open reading frame was introduced using the OS® Site-Directed Mutagenesis Kit (New England Biolabs). For recombinant expression, pET-His6-SUMO-MTA1 (wild-type and mutant) was transformed into SHuffle T7 competent E. co/i (New England Biolabs); pET-His6-SUMO-MTA9 was transformed into Lemo (DE3) competent  E. coli  (New England Biolabs); pET-His6-SUMO-p1 and pET-His6-SUMO-p2 were transformed into BL21(DE3) competent  E. coli  (New England Biolabs). IPTG induction was performed at 16′C overnight. Induced cells were resuspended in 25 ml of lysis buffer B (50 mM Tris pH 7.8, 300 mM NaCl, 5% v/v glycerol, 10 mM imidazole, 5 mM BME, 1 mM PMSF, 0.5× ProBlock Gold Bacterial protease inhibitor cocktail [GoldBio]). The cells were sonicated at 35% amplitude for a total of 4 minutes, with a 10 s off, 10 s cycle using a Model 505 Sonic Dismembrator (Fisherbrand). Lysates were cleared by centrifugation at 30,000 g for 30 min at 4° C., mixed with pre-washed Ni-NTA agarose (Invitrogen), and nutated for 45 min at 4° C. The resin was subsequently washed with lysis buffer and eluted in 50 mM Tris pH 7.8, 300 mM NaCl, 5% v/v glycerol, 400 mM glycerol, 5 mM BME, lx ProBlock Gold bacterial protease inhibitor cocktail [GoldBio]). Eluates were dialyzed into lysis buffer B and then digested with TEV protease (gift from S.H. Sternberg) at 4° C. overnight. The resulting mixture was passed through a fresh batch of Ni-NTA agarose (Invitrogen) to remove cleaved affinity tags. The flow-through containing each recombinant protein was flash frozen and used for all downstream methyltransferase assays. 
     Methyltransferase Assays 
     Generation of DNA and RNA Substrates 
     A 954 bp dsDNA PCR product was used in all assays involving  Tetrahymena  nuclear extract. This substrate was amplified by PCR from  Tetrahymena thermophila  strain SB210 macronuclear SB210 genomic DNA using PCR primers metGATC F2 and metGATC_R2 (Table 2). The resulting product was purified using Ampure XP beads (Beckman Coulter). This 954 bp region of the genome contains a high level of 6 mA in vivo. Thus, the underlying DNA sequence may be intrinsically amenable to methylation by  Tetrahymena  MTA1. Note that the amplified 954 bp product is devoid of DNA methylation as unmodified dNTPs were used for PCR. Separately, a 350 bp dsDNA PCR product was used in all assays involving recombinant MTA1, MTA9, p1 and p2. This sequence lacks 5′-NATC-3′ motifs, and was used to reduce background DNA methylation from contaminating Dam methyltransferase in recombinant protein preparations. The 350 bp dsDNA PCR product was amplified from  Tetrahymena thermophila  strain SB210 macronuclear SB210 genomic DNA using the PCR primers noGATC2 F and noGATC2_R (Table 2), and purified using Ampure XP beads (Beckman Coulter). 
     For short DNA substrates (&lt;50 bp), oligonucleotides were purchased from Integrated DNA Technologies and either directly used as ssDNA, or annealed with its complementary sequence to obtain dsDNA. To prepare hemimethylated 27 bp dsDNA in  FIG. 2G , either strand was methylated using EcoGII methyltransferase (New England BioLabs) before annealing with the complementary sequence. 
     To generate ˜350nt ssRNA and −350 bp dsRNA, the aforementioned 350 bp dsDNA was first PCR-amplified using primers containing T7 overhangs (primer pairs T7noGATC2_F2/noGATC2_R and T7noGATC2_F2/T7noGATC2_R2 respectively; see Table 2 for primer sequences). Each PCR product was used as a template for in vitro transcription using the HiScribe T7 High Yield RNA Synthesis Kit (New England Biolabs). The synthesized RNA was rigorously treated with DNase (ThermoFisher) purified using acid phenol:chloroform extraction, followed by two rounds of chloroform extraction. Each sample was subsequently ethanol precipitated and resuspended in water for use in methyltransferase assays. 
     Radioactive Methyltransferase Assay 
     For experiments involving nuclear extract, 2.18 μg of 954 bp dsDNA substrate was mixed with 4-8 μl nuclear extract and 0.64 μM 3H-labeled S-adenosyl-L-methionine ([ 3 H]SAM) in 33 mM Tris-HCl pH 7.5. 6 mM EDTA. 4.3 mM BME. in a 15p1 reaction volume. For experiments involving recombinant MTA1c protein components (ie. MTA1, MTA9, p1, and/or p2), ˜3 μM oligonucleotide ssDNA/annealed dsDNA is used. Alternatively, 1.3 μg of 350 bp dsDNA substrate (or an equimolar amount ˜350nt ssRNA, or ˜350 bp dsDNA) was used in place of DNA oligonucleotide substrates. ssRNA was heated at 90° C. for 2 min and snap cooled to minimize secondary structures before mixing with other components of the methyltransferase assay. All samples were incubated overnight at 37° C., and subsequently spotted onto 1 cm×1 cm squares of Hybond-XL membrane (GE Healthcare). Membranes were then washed thrice with 0.2M ammonium bicarbonate, once with distilled water, twice with 100% ethanol, and finally air-dried for 1 hr. Each membrane was immersed in 5 mL Ultima Gold (PerkinElmer) and used for scintillation counting on a TriCarb 2910 TR (Perkin Elmer). 
     Non-Radioactive Methyltransferase Assay 
     For assays involving nuclear extract: 5.5 pg of 954 bp DNA substrate was mixed with 20 nuclear extract and 0.2 mM S-adenosyl-L methionine (NEB) in 33 mM Tris-HCl pH 7.5, 6 mM EDTA, 4.3 mM BME in a 15p1 reaction volume. For assays involving recombinant MTA1c protein components (ie. MTA1, MTA9, p1, and/or p2), 2.6 μg of 350 bp DNA substrate was mixed with 540 nM MTA1, 90 nM MTA9, 1.5 μM p1, 1.0 μM p2 proteins. The band of expected size in each recombinant protein preparation was compared against a series of BSA standards to calculate protein concentration. All methylation reactions were incubated at 37° C. overnight, then purified using a MinElute purification kit (QIAGEN), denatured at 95° C. for 10 min, and snap cooled in an ice water bath. Samples were spotted on a Hybond N+ membrane (GE Healthcare), air-dried for 5 min and UV-cross-linked with 120,000 μJ/cm 2  exposure using an Ultra-Lum UVC-515 Ultraviolet Multilinker. The cross-linked membrane was blocked in 5% milk in TBST (containing 0.1% v/v Tween) and incubated with 1:1,000 anti-N6-methyladenosine antibody (Synaptic Systems) at 4° C. overnight. The membrane was then washed three times with TBST, incubated with 1:3,000 Goat anti-rabbit HRP antibody (Bio-Rad) at room temperature for 1 hr, washed another three times with 1×TBST, and developed using Amersham ECL Western Blotting Detection Kit (GE Healthcare). This dot blot assay was used to measure 6 mA levels in  FIGS. 2F, 3B, 5C, and 10C . 
     Quantitative Mass Spectrometry Analysis of dA and 6 mA 
     10.5 μg  Oxytricha  or  Tetrahymena  macronuclear genomic DNA was first digested to nucleosides by mixing with 14p1 DNA degradase plus enzyme (Zymo Research) in a 262.5 μl reaction volume. Samples were incubated at 37° C. overnight, then 70° C. for 20 min to deactivate the enzyme. 
     The internal nucleoside standards  15 N 5 -dA and D 3 -6 mA were used to quantify endogenous dA and 6 mA levels in ciliate DNA.  15 N 5 -dA was purchased from Cambridge Isotope Laboratories, while D 3 -6 mA was synthesized as described in the following section. Nucleoside samples were spiked with 1 ng/μl  15 N 5 -dA and 200 pg/μl D 3 -6 mA in an autosampler vial. Samples were loaded onto a 1 mm×100 mm C18 column (Ace C18-AR, Mac-Mod) using a Shimadzu HPLC system and PAL auto-sampler (20 μl/injection) at a flow rate of 70 μl/min. The column was connected inline to an electrospray source couple to an LTQ-Orbitrap XL mass spectrometer (Thermo Fisher). Caffeine (2 pmol/μl in 50% Acetonitrile with 0.1% FA) was injected as a lock mass through a tee at the column outlet using a syringe pump at 0.5p1/min (Harvard PHD 2000). Chromatographic separation was achieved with a linear gradient from 10% to 99% B (A: 0.1% Formic Acid, B: 0.1% Formic Acid in Acetonitrile) in 5 min, followed by 5 min wash at 100% B and equilibration for 10 min with 1% B (total 20 min program). Electrospray ionization was achieved using a spray voltage of 4.50 kV aided by sheath gas (Nitrogen) flow rate of 18 (arbitrary units) and auxiliary gas (Nitrogen) flow rate of 2 (arbitrary units). Full scan MS data were acquired in the Orbitrap at a resolution of 60,000 in profile mode from the m/z range of 190-290. A parent mass list was utilized to acquire MS/MS spectra at a resolution of 7500 in the Orbitrap. LC-MS data were manually interpreted in Xcalibur&#39;s Qual browser (Thermo, Version 2.1) to visualize nucleoside mass spectra and to generate extracted ion chromatograms by using the theoretical [M+H] within a range of ±2 ppm. Peak areas were extracted in Skyline (Ver. 3.5.0.9319). 
     Synthesis of D 3 -6 mA Nucleoside 
     2′-Deoxyadenosine and CD3I were purchased from Sigma Aldrich. Flash chromatography was performed on a Biotage Isolera using silica columns (Biotage SNAP Ultra, HP-Sphere 25 pm). Semi-preparative RP-HPLC was performed on a Hewlett-Packard 1200 series instrument equipped with a Waters XBridge BEH C18 column (5 μm, 10×250 mm) at a flow rate of 4 mL/min, eluting using A (0.1% formic acid in H 2 O) and B (0.1% formic acid in 9:1 MeCN/H 2 O).  1 H NMR spectra were recorded on a Bruker UltraShield Plus 500 MHz instrument. Data for  1 H NMR are reported as follows: chemical shift (8 ppm), multiplicity (s=singlet, br=broad signal, d=doublet, dd=doublet of doublets) and coupling constant (Hz) where possible.  13 C NMR spectra were recorded on a Bruker UltraShield Plus 500 MHz. 
     D 3 -6 mA (2′Deoxy-6-[D3]-methyladenosine) were synthesized and purified according to (Schiffers et al., 2017). After an initial purification by flash column chromatography, the methylated compounds were further purified by semipreparative RP-HPLC (linear gradient of 0% to 20% B over 30 min) affording the desired compounds in 14% and 10% yields respectively after lyophilization. 
     2Deoxy-6-[D3]methyladenosine 
       1 H NMR (500 MHz, D 2 O) δ 7.98 (s, 1H), 7.77 (s, 1H), 6.17 (m, 1H), 4.54 (m, 1H), 4.10 (m, 1H), 3.79 (dd, J=12.7, 3.2 Hz, 1H), 3.71 (dd, J=12.7, 4.3 Hz, 1H), 2.60 (m, 1H), 2.44 (ddd, J=14.0, 6.3, 3.3 Hz, 1H). 
       13 C NMR (126 MHz, D 2 O) δ 154.0, 151.5, 146.1, 138.9, 118.4, 87.3, 84.3, 71.1, 61.6, 39.2, 26.4 ppm. (Peak at 26.4 ppm appears as a broad signal. C-D coupling is not resolved). 
     HR-MS (ESI+): m/z calculated for [C 11 H 13 D 3 N 5 O 3 ] +  ([M+Hr): 269.1436. found 269.1421. 
     Mass Spectrometry Analysis of Proteins in  Tetrahymena  Nuclear Extracts 
     Samples where topped up to 200p1 with 50 mM ammonium bicarbonate pH 8. TCEP was added to 5 mM final concentration and left to incubate at 60° C. for 10 min. 15 mM chloroacetamide was then added and left to incubate in the dark at room temperature for 30 min. 1 μg of Trypsin Gold (Promega) was added to each sample and incubated end-over-end at 37° C. for 16 hr. An additional 0.25 μg of Trypsin Gold was added and incubated end-over-end at 37° C. for 3 hr. Samples were acidified by adding TFA to 0.2% final concentration, and desalted using SDB stage-tips (Rappsilber et al., 2007). Samples were dried completely in a speedvac and resuspended in 20p1 of 0.1% formic acid pH 3.5 μl was injected per run using an Easy-nLC 1200 UPLC system. Samples were loaded directly onto a 45 cm long 75 pm inner diameter nano capillary column packed with 1.9 μm C18-AQ (Dr. Maisch, Germany) mated to metal emitter in-line with an Orbitrap Fusion Lumos (Thermo Scientific, USA). The mass spectrometer was operated in data dependent mode with the 120,000 resolution MS1 scan (AGC 4e5, Max IT 50 ms, 400-1500 m/z) in the Orbitrap followed by up to 20 MS/MS scans with CID fragmentation in the ion trap. Dynamic exclusion list was invoked to exclude previously sequenced peptides for 60 s if sequenced within the last 30 s, and maximum cycle time of 3 s was used. Peptides were isolated for fragmentation using the quadrupole (1.6 Da window). Ns was utilized. Ion-trap was operated in Rapid mode with AGC 2e3, maximum IT of 300 msec and minimum of 5000 ions. 
     Raw files were searched using Byonic (Bern et al., 2012) and Sequest HT algorithms (Eng et al., 1994) within the Proteome Discoverer 2.1 suite (Thermo Scientific, USA). 1 Oppm MS1 and 0.4 Da MS2 mass tolerances were specified. Caramidomethylation of cysteine was used as fixed modification, while oxidation of methionine, pyro-Glu from Gln and deamidation of asparagine were specified as dynamic modifications. Trypsin digestion with maximum of 2 missed cleavages were allowed. Files were searched against the  Tetrahymena themophila  macronuclear reference proteome (June 2014 build), supplemented with common contaminants (27,099 total entries). 
     Scaffold (version Scaffold 4.8.7, Proteome Software Inc., Portland, Oreg.) was used to validate MS/MS based peptide and protein identifications. Peptide identifications were accepted if they could be established at greater than 93.0% probability. Peptide Probabilities from Sequest and Byonic were assigned by the Scaffold Local FDR algorithm. Protein identifications were accepted if they could be established at greater than 99.0% probability to achieve an FDR less than 1.0% and contained at least 3 identified peptides. Protein probabilities were assigned by the Protein Prophet algorithm (Nesvizhskii et al., 2003). Proteins that contained similar peptides and could not be differentiated based on MS/MS analysis alone were grouped to satisfy the principles of parsimony. 
     Generation of Mta1 Mutant Lines 
     A frameshift mutation in the MTA1 gene was created by inserting a small non-coding DNA segment immediately downstream of the MTA1 start codon ( FIGS. 3A and 12H ). This non-coding DNA segment belongs to a class of genetic elements that are normally eliminated during the sexual cycle (Chen et al., 2014). When ssRNA homologous to such DNA segments is injected into  Oxytricha  cells undergoing sexual development, the DNA is erroneously retained (Khurana et al., 2018). This results in disruption of the MTA1 open reading frame. The ectopic DNA segment is propagated through subsequent cell divisions after completion of the sexual cycle. RNaseq analysis confirmed the presence of the ectopic insertion in mta1 mutant transcripts but not wild-type controls ( FIG. 12H ). 
     ssRNA was generated by in vitro transcription using a Hi-Scribe T7 High Yield RNA Synthesis Kit (New England Biolabs). The DNA template for in vitro transcription consists of the ectopic DNA segment flanked by 100-200 bp cognate MTA1 sequence. Following DNase treatment, ssRNA was acid-phenol:chloroform extracted and ethanol precipitated. After precipitation, ssRNA was resuspended in nuclease-free water (Ambion) to a final concentration of 1 to 3 mg/mL for injection. 
     ssRNA Microinjections 
       Oxytricha  cells were mated by mixing 3 mL of each mating type, JRB310 and JRB510, along with 6 mL of fresh Pringsheim media. At 10 to 12 hr post mixing, pairs were isolated and placed in Volvic water with 0.2% bovine serum albumin (Jackson ImmunoResearch Laboratories) (Fang et al., 2012). ssRNA constructs were injected into the macronuclei of paired cells under a light microscope as previously described with DNA constructs (Nowacki et al., 2008). After injection, cells were pooled in Volvic water. At 60 to 72 hr post mixing, the pooled cells were singled out to grow clonal injected cell lines. As clonal population size grew, lines were transferred to 10 cm Petri dishes and grown in Pringsheim media. Only water from the “Volvic” brand has been empirically tested in our laboratory to support  Oxytricha  growth. Similar products from other vendors have not been tested. 
     Survival Analysis of  Oxytricha  Mta1 Mutants 
     This experiment was performed in  FIG. 7D . Wild-type or mutant  Oxytricha  cells were mixed at 0 hr to induce mating. Since not all cells enter the sexual cycle, mated cells are separated from unmated vegetative cells at 15 hr and transferred into a separate dish. The cells are allowed to rest for 12 hr to account for cell death during transfer. The number of surviving mated cells is counted from 27 hr onward. The total cell number at each time point is normalized to 27 hr data to obtain the percentage survival. An increase in survival at 108 hr is observed in wild-type samples because the cells have completed mating and reverted to the vegetative state, where they can proliferate and increase in number. 
     Quantification and Statistical Analysis 
     All statistical tests were performed in Python (v2.7.10) or R (v3.2.5), and described in the respective Figure and Table legends. 
     Data and Software Availability 
       Oxytricha  SMRT-seq data are deposited in SRA under the accession numbers SRA: SRX2335608 and SRX2335607, and GEO: GSE94421 . Tetrahymena  SMRT-seq and all  Oxytricha  Illumina data are deposited in NCBI GEO under accession number GEO: GSE94421. 
     
       
         
           
               
             
               
                 TABLE 1 
               
               
                   
               
               
                 Protein sequences for phylogenetic tree construction. 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
            
               
                 Protein sequences for phylogenetic analysis of MT-A70 proteins (including MTA1 and MTA9) 
               
               
                 &gt;NP_495127.1 DNA N6-methyl methyltransferase [ Caenorhabditis elegans ] 
               
               
                 (SEQ ID No: 1) 
               
               
                 MDTEFAILDEEKYYDSVFKELNLKTRSELYEISSKFMPDSQFEAIKRRGISNRKRKIKETSENSNRMEQMALKIKNVG 
               
               
                   
               
               
                 TELKIFKKKSILDNNLKSRKAAETALNVSIPSASASSEQIIEFQKSESLSNLMSNGMINNWVRCSGDKPGIIENSDGTK 
               
               
                   
               
               
                 FYIPPKSTFHVGDVKDIEQYSRAHDLLFDLIIADPPWFSKSVKRKRTYQMDEEVLDCLDIPVILTHDALIAFWITNRIGI 
               
               
                   
               
               
                 EEEMIERFDKWGMEVVATWKLLKITTQGDPVYDFDNQKHKVPFESLMLAKKKDSMRKFELPENFVFASVPMSVHS 
               
               
                   
               
               
                 HKPPLLDLLRHFGIEFTEPLELFARSLLPSTHSVGYEPFLLQSEHVFTRNISL 
               
               
                   
               
               
                 &gt;NP_564080.1 Methyltransferase MT-A70 family protein [ Arabidopsis thaliana ] 
               
               
                 (SEQ ID No: 2) 
               
               
                 MAKTDKLAQFLDSGIYESDEFNWFFLDTVRITNRSYTRFKVSPSAYYSRFFNSKQLNQHSSESNPKKRKRKQKNSS 
               
               
                   
               
               
                 FHLPSVGEQASNLRHQEARLFLSKAHESFLKEIELLSLTKGLSDDNDDDDSSLLNKCCDDEVSFIELGGVWQAPFYE 
               
               
                   
               
               
                 ITLSFNLHCDNEGESCNEQRVFQVFNNLVVNEIGEEVEAEFSNRRYIMPRNSCFYMSDLHHIRNLVPAKSEEGYNLI 
               
               
                   
               
               
                 VIDPPWENASAHQKSKYPTLPNQYFLSLPIKQLAHAEGALVALWVTNREKLLSFVEKELFPAWGIKYVATMYWLKV 
               
               
                   
               
               
                 KPDGTLICDLDLVHHKPYEYLLLGYHFTELAGSEKRSDFKLLDKNQIIMSIPGDFSRKPPIGDILLKHTPGSQPARCLE 
               
               
                   
               
               
                 &gt;ORY94237.1 MT-A70-domain-containing protein [ Syncephalastrum racemosum ] 
               
               
                 (SEQ ID No: 3) 
               
               
                 MIVASSDTCDIVDCEAAFGIDGTVRLRPGDFSLGTPYFTSRLGQKRPRPDDDTLDNTPSDTIHAIVQQLPVMAPDY 
               
               
                   
               
               
                 WHDRPMEAVVMNAHVHFPSLVSLAEASLRFDPDNDEDEDNRQILRPDMALESLQVFYRHFEHPKDSPILIRVQDAY 
               
               
                   
               
               
                 YWIPPRTAFMMGSLENIHLPTLGKFDCIVMDPPWPNKSVRRSAHYETQEDIYDLFAIPLPQLAQPNCLVAVWVTNK 
               
               
                   
               
               
                 PKFIRFVQKLFAAWDVEPLTTWYWLKVTTHGEPVCPIDSPHRKPYEHLILGRKRPVKININDPPALPRVLVSVPSKH 
               
               
                   
               
               
                 HSRKPPLNDILMRYLPSDARRLELFARCLTPGWTSWGNECLKFQHVDYFYDTNEAMEEGKQK 
               
               
                   
               
               
                 &gt;ORX58127.1 MT-A70-domain-containing protein [ Hesseltinella vesiculosa ] 
               
               
                 (SEQ ID No: 4) 
               
               
                 MANAARRFAQQDELPLDVSQDLQDLPLLDLFNRKVINDSDQCSSLHVASFGQYLVPRHTKFVMSDLDNIDLLRSEN 
               
               
                   
               
               
                 DVFDLIVMDPPWPNKSVHRSTDYETQDIYDLFHLPIKSLIKNQGLVAVWVTNKPKYRRFILDKLFKAWQMTCVGEW 
               
               
                   
               
               
                 LWLKVTSSGEPVFPLDSPHRKPYEQLILGRYQPDDTSPTLPNPPQQHVLISVPSIRHSRKPPLGEVLADFLPKQPAC 
               
               
                   
               
               
                 LELFARCLTPGWTSWGNECLKFQHESYFISNDTPHSPSAS 
               
               
                   
               
               
                 &gt;ORZ15132.1 MT-A70-like protein, partial [ Absidia repens ] 
               
               
                 (SEQ ID No: 5) 
               
               
                 YDLVVMDPPWPNKSVHRSSHYETQDIYDLYQIPLTSLVHKNSLVAVWITNKPKYRRFVMDKLFKSWHVDCVAEWT 
               
               
                   
               
               
                 WLKVTNDGEPVFPLNSTHRKPYEQLIIGRYNGGSGGGNDNNDSIQEESEVKPIPYQHSIVSVPSKRHSRKPPLQDL 
               
               
                   
               
               
                 LQPYLPAKPRCLELFARCLTPGWSSWGNECLKFQNEYYYTRIENPLHIDRSDV 
               
               
                   
               
               
                 &gt;XP_021679935.1 MT-A70-domain-containing protein [ Lobosporangium transversale ] 
               
               
                 (SEQ ID No: 6) 
               
               
                 MLHESTVSVLDRLILISHISLQTYLLAKDREGFDIIVMDPPWQNASVDRMSHYRTMDLYELFKIPIPDLLKANGSNVG 
               
               
                   
               
               
                 GIVAVWITNKAKVKRVVVEKLFPAWGLDLVAHWFWLKVTTKGEPVLSLSNSHRRAYEGVLIGRQRQGSKLSNKTM 
               
               
                   
               
               
                 HETSASNPVNRLLVSIPAQHSRKPSLNALIEEEFFTSKLESRADRDRNAYVDSEALVKKPLYRLELFARNLEEGVLS 
               
               
                   
               
               
                 WGNEPLRYQYCGRGASNSQVVQDGYLIPCPIQSELVSQ 
               
               
                   
               
               
                 &gt;XP_689178.3 methyltransferase-like protein 4 isoform X1 [ Danio rerio ] 
               
               
                 (SEQ ID No: 7) 
               
               
                 MSVVCCNSWGWLLDSSSHIDKDFQRCVCYNEANGLEENTHFTCCFKRQYFNILMPHMQQSTAMSGFPLDSGKH 
               
               
                   
               
               
                 DSAEHEKIELQTRKKRKRKHHDLNTGEIEANIYHDKVRSVVLEGSRALLEAGRQCGYFTEALTESQTISTPSESTSA 
               
               
                   
               
               
                 HECQLAAFCDLAKQLPLSEESPVHTLSRDGQNPALDLFSSITENPFDCACEITFMRERYLLPPRCRFLLSDVTRMDP 
               
               
                   
               
               
                 LVNSGDKFDLIVLDPPWENKSVKRSNRYSSLPSSQLKKLPVPALAAPGGLVVTWVTNRAKHRRFVREELYPHWAV 
               
               
                   
               
               
                 EVLAEWLWVKVTRSGEFVFPLDSQHKKPYEVLVLGRCRSTSDHTDRCSAVNELPDQRLLVSVPSTLHSHKPSLAA 
               
               
                   
               
               
                 VLKPYIRREPRCLELFARSLQSDWSCWGNEVLKFQHCSYFSRHTDQEPTSDTLQRTHSHLQSTGLLETPETAR 
               
               
                   
               
               
                 &gt;NP_073751.3 methyltransferase-like protein 4 isoform 1 [ Homo sapiens ] 
               
               
                 (SEQ ID No: 8) 
               
               
                 MSVVHQLSAGWLLDHLSFINKINYQLHQHHEPCCRKKEFTTSVHFESLQMDSVSSSGVCAAFIASDSSTKPENDDG 
               
               
                   
               
               
                 GNYEMFTRKFVFRPELFDVTKPYITPAVHKECQQSNEKEDLMNGVKKEISISIIGKKRKRCVVFNQGELDAMEYHTKI 
               
               
                   
               
               
                 RELILDGSLQLIQEGLKSGFLYPLFEKQDKGSKPITLPLDACSLSELCEMAKHLPSLNEMEHQTLQLVEEDTSVTEQD 
               
               
                   
               
               
                 LFLRVVENNSSFTKVITLMGQKYLLPPKSSFLLSDISCMQPLLNYRKTFDVIVIDPPWQNKSVKRSNRYSYLSPLQIQ 
               
               
                   
               
               
                 QIPIPKLAAPNCLLVTWVTNRQKHLRFIKEELYPSWSVEVVAEWHWVKITNSGEFVFPLDSPHKKPYEGLILGRVQE 
               
               
                   
               
               
                 KTALPLRNADVNVLPIPDHKLIVSVPCTLHSHKPPLAEVLKDYIKPDGEYLELFARNLQPGWTSWGNEVLKFQHVDY 
               
               
                   
               
               
                 FIAVESGS 
               
               
                   
               
               
                 &gt;XP_020951799.1 methyltransferase-like protein 4 isoform X1 [ Sus scrofa ] 
               
               
                 (SEQ ID No: 9) 
               
               
                 MSVVHQLSSGWLLDHLSFINKISYELHQHHEPCCSKNEPTSVHLDSLHKDSVFSFGASPAFIASSSKPENDDGGNR 
               
               
                   
               
               
                 EMSMQKYVFRSELFDVTKPYITSAIHKECQQSNEKEDLANDVKKEASISIKRKKRKRCVVFNQGELDAMEYHTKIRG 
               
               
                   
               
               
                 LILDGSSQLIQEGLKSGFLHPLSEKCDKCSKPVTLPLDTCSLSELCEMAKHVPSLNEMELQTLQLMEDDISVTEQDLF 
               
               
                   
               
               
                 SRIVENNSSFTKMITLMGQKYLLPPKSSFLLSDISCIYPLLNCRKTYDVIVIDPPWQNKSVKRSNRYSYLSPLQIKQIPI 
               
               
                   
               
               
                 PKLAAPNCLVVTWVTNRQKHLRFVKEELYPSWSVEIVAEWHWVKITNSGEFVFPIDSPHKKPYEVLVLGRVRERAA 
               
               
                   
               
               
                 LLLSRNAEVKELSIPDRKLIVSVPCILHSHKPPLAEVLKDYIKPEGEYLELFARNLQPGWTSWGNEVLKFQHMDYFVA 
               
               
                   
               
               
                 LESRS 
               
               
                   
               
               
                 &gt;XP_011245012.1 PREDICTED: methyltransferase-like protein 4 isoform X2 [ Mus musculus ] 
               
               
                 (SEQ ID No: 10) 
               
               
                 MSVVHHLPPGWLLDHLSFINKVNYQLCQHQESFCSKNNPTSSVYMDSLQLDPGSPFGAPAMCFAPDFTTVSGND 
               
               
                   
               
               
                 DEGSCEVITEKYVFRSELFNVTKPYIVPAVHKERQQSNKNENLVTDYKQEVSVSVGKKRKRCIAFNQGELDAMEYH 
               
               
                   
               
               
                 TKIRELILDGSSKLIQEGLRSGFLYPLVEKQDGSSGCITLPLDACNLSELCEMAKHLPSLNEMELQTLQLMGDDVSVI 
               
               
                   
               
               
                 ELDLSSQIIENNSSFSKMITLMGQKYLLPPQSSFLLSDISCMQPLLNCGKTFDAIVIDPPWENKSVKRSNRYSSLSPQ 
               
               
                   
               
               
                 QIKRMPIPKLAAADCLIVTWVTNRQKHLCFVKEELYPSWSVEVVAEWYWVKITNSGEFVFPLDSPHKKPYECLVLG 
               
               
                   
               
               
                 RVKEKTPLALRNPDVRIPPVPDQKLIVSVPCVLHSHKPPLTGYLNSSFATLIPRVSNNMEYCRVVRTAFIA 
               
               
                   
               
               
                 &gt;XP_018079135.1 PREDICTED: methyltransferase-like protein 4 [ Xenopus laevis ] 
               
               
                 (SEQ ID No: 11) 
               
               
                 MSVVCETSAGWLVDELSLLRKWYQHSTSCQDAAHKKQLYDIKEDLFLILRPHIPVQSTPAPLPILCPETNPGTINQR 
               
               
                   
               
               
                 KKRKRSCAFNQGELDAMEYHKKIIDFIMEGTQPLLQEGFKRLFLRPVLVNDDDHSQTEPRLCNNPCQLAELCNMAK 
               
               
                   
               
               
                 CMPLLNPGEHAVQVLERGIYLPQETNVLSCITENKSECPEVIQFMGEKYIIPPKSTFLMSDVSCMEPLLHYKRYNIIVM 
               
               
                   
               
               
                 DPPWENKSVKRSKRYSSLSPNEIQQLPVPVLAAPDCLVITWVTNKQKHLRFVKEDLYPHWSVKTLGEWHWVKITR 
               
               
                   
               
               
                 SGEFVFPLDSTHKKPYEVLIIGRFKGAGNSTARKSEICLPPIPERKLIVSVPCKLHSHKPPLSEILKEYVKPDLECLELF 
               
               
                   
               
               
                 ARNLQPGWTSWGNEVLKFQHIDYFTPVDVED 
               
               
                   
               
               
                 &gt;NP_650573.1 uncharacterized protein Dmel_CG14906 [ Drosophila melanogaster ] 
               
               
                 (SEQ ID No: 12) 
               
               
                 MLKLQKKTEDSKFAVFLDHKTLINEAYDEFKLKSELFQFHAKKTDKGIEEDKTRKRKRKAGVEDASSLEDLHLVNEY 
               
               
                   
               
               
                 LELLSKPVEPEDSSPMKRHWEDGYNVPQLHGANESGRMQRFLRVDGSRGVYLIPNQSRFFNHNVDNLPALLHQLL 
               
               
                   
               
               
                 PAYDLIVLDPPWRNKYIRRLKRAKPELGYSMLSNEQLSHIPLSKLTHPRSLVAIWCTNSTLHQLALEQQLLPSWNLR 
               
               
                   
               
               
                 LLHKLRWYKLSTDHELIAPPQSDLTQKQPYEMLYVACRSDASENYGKDIQQTELIFSVPSIVHSHKPPLLSWLREHLL 
               
               
                   
               
               
                 LDKDQLEPNCLELFARYLHPHFTSIGLEVLKLMDERLYEVRKVEHCNQEEVN 
               
               
                   
               
               
                 &gt;tr|A8J2E1|A8J2E1_CHLRE Predicted protein OS =  Chlamydomonas reinhardtii  OX = 3055 
               
               
                 GN = CHLREDRAFT_174824 PE = 3 SV = 1 
               
               
                 (SEQ ID No: 13) 
               
               
                 MATLPGAAAAAPGANAEVGVPEPSLEPQDALQQRIALAEGLLALNEADAMQAWQQLPREALLEQVAKYRGAVRD 
               
               
                   
               
               
                 MASALRSSTLPGGVPPHCVPIHANVTTFDWPSLYSHAQFDVIMMDPPWQLATANPTRGVALGYSQLNDDHISRLP 
               
               
                   
               
               
                 VPQLQRQGGYLFVWVINAKYKWTLDLFDRWGYRLVDEVVWVKMTVNRRLAKSHGYYLQHAKEVCLVAKRGNPP 
               
               
                   
               
               
                 VPPGCEGGVGSDIIFSERRGQSQKPEEIYHLIEQLVPNGRYLEIFARKNNLRNYWVSIGNEVTGTGLPDEDMQALRD 
               
               
                   
               
               
                 LHHIPGAVYGKNAPHLVSKLFLYAPNSSREEG 
               
               
                   
               
               
                 &gt;XP_021880122.1 MT-A70-domain-containing protein [ Lobosporangium transversale ] 
               
               
                 (SEQ ID No: 14) 
               
               
                 MLDQINIDIEQLEASLDIDEGKAHSNNASGTGCLIGTGTSSGNASNGAGVADEDLEEEVDDLEEFEAPEWCVPIKAN 
               
               
                   
               
               
                 VMTYDWDSLAAECQFDVILMDPPWQLATHAPTRGVAIAYQQLPDICIEELPVPKLSSNGFIFIWVINNKYAKAFDLM 
               
               
                   
               
               
                 RRWGYSYVDDITWVKQTVNRRMAKGHGYYLQHAKETCLVGKKGEDPPGCRHSIGSDVIFSERRGQSQKPEELYE 
               
               
                   
               
               
                 LIEELVPNGRYLEIFGRKNNLRDYWVTVGNEL 
               
               
                   
               
               
                 &gt;ORX69627.1 MT-A70-domain-containing protein [ Linderina pennispora ] 
               
               
                 (SEQ ID No: 15) 
               
               
                 MDVDSSSPAVVLQALRQREQKIRSRILVLEQEISDLEKRCGVEGSGDAANKVTEADLEEFKAPEWSVPIRANVMNF 
               
               
                   
               
               
                 DWEKLAQACQFDVILMDPPWQLASQAPTRGVAIAYQQLPDVCIESLPIDLLQTSGFIFIWVINNKYTKAFQLMKQWG 
               
               
                   
               
               
                 YKYVDDIAWVKQTVNRRMAKGHGYYLQHAKETCLVGKKGPDPPNLRRSVASDVIFSERRGQSQKPEELYEIIEQLV 
               
               
                   
               
               
                 PGGRYLEIFGRKNNLRDYWVTVGNEL 
               
               
                   
               
               
                 &gt;ORX98979.1 allantoinase [ Basidiobolus meristosporus  CBS 931.73] 
               
               
                 (SEQ ID No: 16) 
               
               
                 MSAIIFTGNRVLFDSTSKVEPATIHVDPWTGRIVKITNKRSTKADFPGIEDKDFVDAGDDLIMPGVIDAHVHLNEPGR 
               
               
                   
               
               
                 TDWEGFDTATRAAAAGGLTTVIDMPLNSIPPTTTLENLNTKKEAAKPQAWVDVGFYGGVIPGNADQLRPMIAAGVC 
               
               
                   
               
               
                 GFKCFLIESGVDEFPCVNEEEVRKAFAEFDGTDNVFMFHAEMECDDHSHETAAPQSTDPSAYQTFLQSRPHALEV 
               
               
                   
               
               
                 KAIEMIIRVCKDFPNVRAHIVHLSSAEALPMIRKAKAEGVKLTVETCYHYLTLNAEDIINGATHFKCCPPIREGSNRELL 
               
               
                   
               
               
                 WEALLDGTIDYVVSDHSPCTPELKRFDSGDFTAAWGGISSLQFGLSLLWTEAKRRGCTLQDLTRWLSQNTARHAG 
               
               
                   
               
               
                 ILNRKGRLQIGSDADIVIWSPEETFVVDKKMIHFKNKVTPYENMTLHGAVKKTFVRGRNVYDKSTAQLFSAKPLGNL 
               
               
                   
               
               
                 LARFQVYSNPITAMPSYAQPPSSDNGDFEEESEDYIESDEVDEDLRELLAKETSLRLRIDSLKEEILKLEREQRGETD 
               
               
                   
               
               
                 GSKNEGEGGEEEIDLEEFEAPEWCVPIKANVMTFEWKRLAEAAQFDVILMDPPWQLATHAPTRGVAIGYQQLPDV 
               
               
                   
               
               
                 CIEELPIPLLQKNGFIFIWVINNKYVKAFELMAKWGYRYVDDITWVKQTVNRRMAKGHGYYLQHAKETCLIGKKGED 
               
               
                   
               
               
                 PPNCRHSVCSDVIFSERRGQSQKPEELYEMIEQLVPNGKYLEIFGRKNNLRDYWVTIGNEL 
               
               
                   
               
               
                 &gt;ORZ00623.1 MT-A70-domain-containing protein [ Syncephalastrum racemosum ] 
               
               
                 (SEQ ID No: 17) 
               
               
                 MSSREESPSSVSGFDLDTIDESTVTDTTLKNLLRREIELQLQIDALQTEILQIEESTAAGKNNKNDEELDPQDLEEFEA 
               
               
                   
               
               
                 PEWCVPIKANVMTFDWEALASEVQFDVIVADPPWQLATHAPTRGVAIGYQQLPDVCIEEIPIQKLQKNGFIFIWVINN 
               
               
                   
               
               
                 KYAKAFELMERWGYHYVDDITWVKQTVNRRMAKGHGYYLQHAKETCLVGKKGEDPPNCRHSVGSDVIFSERRG 
               
               
                   
               
               
                 QSQKPEELYELIEELVPNGKYLEIFGRKNNLRDYWVTVGNEL 
               
               
                   
               
               
                 &gt;ORZ06213.1 MT-A70-domain-containing protein [ Absidia repens ] 
               
               
                 (SEQ ID No: 18) 
               
               
                 MTSDTSAMTADVLNRKRKRSPAMNGDDLSNNSDEADNNTTTGTTTSVDSNENDYQEQDREPILRLPRLNDAKLLE 
               
               
                   
               
               
                 EVVDDVDYEDQPERYDFDFKKLWLQERGLMERIDGLLKDIARLTDFKGHYRDMVIPSDDEDDLDDEDSKAQYDAP 
               
               
                   
               
               
                 EWCVPIKANVMTFDWESLGKEVQFDVIMADPPWQLATHAPTRGVAISYQQLPDVCIEDLPLEKLQTNGFLFIWVIN 
               
               
                   
               
               
                 NKYAKAFEMMEKWGYKYVDDITWVKQTVNRRMAKGHGYYLQHAKETCLVGVKGTLPPYCRRSVGSDVIYSERRG 
               
               
                   
               
               
                 QSQKPEQIYELIEEMMPGGKYLEIFGRKNNLRDYWITVGNEL 
               
               
                   
               
               
                 &gt;ORX43344.1 MT-A70-domain-containing protein [ Hesseltinella vesiculosa ] 
               
               
                 (SEQ ID No: 19) 
               
               
                 MASESNISRESSPASISSTNSESGIENVQSLTDEDLKQLILKEMNLKEHIEQLQRKISKLTANDLSTNQDSSDADDDLL 
               
               
                   
               
               
                 NGDETMDDDSSSGSDSEVSGNEDIASVKSSPHAADKSESESESESDEGSSEDGNDEEDEFEAPKWCVPIKANVM 
               
               
                   
               
               
                 TFDWEKLASETQFDVIVADPPWQLATHAPTRGVAIAYQQLPDVCIEDLPIEKLQTNGFIFIWVINNKYAKAFELMEKM 
               
               
                   
               
               
                 GYTYVDDITWVKQTVNRRMAKGHGYYLQHAKETCLVGKKGVDPPSCRHSVGSDIIFSERRGQSQKPEELYELIEEL 
               
               
                   
               
               
                 VPNGKYLEIFGRKNNLRDYWVTVGNEL 
               
               
                   
               
               
                 &gt;ORX52920.1 MT-A70 protein [ Piromyces finnis ] 
               
               
                 (SEQ ID No: 20) 
               
               
                 MMIVANEIDYEEFTAPEWCIPIKANVIDFEWDKLASECQFDAILMDPPWQLATHAPTRGVAIAYQQLPDQFIEELPIE 
               
               
                   
               
               
                 KLQKNGFIFIWVINNKYVKAFELMKKWGYTFVDDITWVKQTVNRRMAKGHGYYLQHAKETCLVGKKGEDPVGCKH 
               
               
                   
               
               
                 SISSDVIYSVRRGQSQKPEELYEMIEELIPNGKYLEIFGRKNNLRDYWVTIGNEL 
               
               
                   
               
               
                 &gt;ORX86973.1 MT-A70-domain-containing protein [ Anaeromyces robustus ] 
               
               
                 (SEQ ID No: 21) 
               
               
                 MDEKEVENSVLDSSNIEKSNATTSNMDVDETSNNETSTAIIKSEDGANSYDDFLKLDFTPEEEKDEVLKKLIERETEL 
               
               
                   
               
               
                 KLKIEKEIEGIKNLELKGFSALTQKDEDVQDIDYEEFTAPEWCIPIKANVIDFEWDKLASECQFDAILMDPPWQLATHA 
               
               
                   
               
               
                 PTRGVAIAYQQLPDQFIEELPIEKLQKNGFIFIWVINNKYVKAFELMKKWGYTFVDDITWVKQTVNRRMAKGHGYYL 
               
               
                   
               
               
                 QHAKETCLVGKKGDDPVGCRHKISSDVIYSVRRGQSQKPEELYEMIEELIPNGKYLEIFGRKNNLRDYWVTIGNEL 
               
               
                   
               
               
                 &gt;XP_001032074.3 MT-a70 family protein [ Tetrahymena thermophila  SB210] 
               
               
                 (SEQ ID No: 22) 
               
               
                 MKKEQQFLIFKKSLIIAQKRKEINIKQLKQQFKNFLFVQIFSIIKLKLQDIIIKFKMSKAVNKKGLRPRKSDSILDHIKNKLD 
               
               
                   
               
               
                 QEFLEDNENGEQSDEDYDQKSLNKAKKPYKKRQTQNGSELVISQQKTKAKASANNKKSAKNSQKLDEEEKIVEEE 
               
               
                   
               
               
                 DLSPQKNGAVSEDDQQQEASTQEDDYLDRLPKSKKGLQGLLQDIEKRILHYKQLFFKEQNEIANGKRSMVPDNSIPI 
               
               
                   
               
               
                 CSDVTKLNFQALIDAQMRHAGKMFDVIMMDPPWQLSSSQPSRGVAIAYDSLSDEKIQNMPIQSLQQDGFIFVWAIN 
               
               
                   
               
               
                 AKYRVTIKMIENWGYKLVDEITWVKKTVNGKIAKGHGFYLQHAKESCLIGVKGDVDNGRFKKNIASDVIFSERRGQS 
               
               
                   
               
               
                 QKPEEIYQYINQLCPNGNYLEIFARRNNLHDNWVSIGNEL 
               
               
                   
               
               
                 &gt;EJY88228.1 MT-A70 family protein [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 23) 
               
               
                 MNQSSQDITTQKSSNGFNPQTQPETLIQVIRKESTFIFKYRKNPYYVPPPISSQTSPNLEVETSNDLNQMSDYEGQI 
               
               
                   
               
               
                 PNNYEINRNSTQFTNNDDQSDNDFYDNNSITTMQIDTSTAKILNNGPLEYNPDLPNKEQKLKDSQVMQNQPPTATS 
               
               
                   
               
               
                 TNSQQRTLQELINIMPSIEDISQQCKQQQQLKIQAKANSTQSASTANAANGGKGRKRGRTVRFDQPLLGKVRQRN 
               
               
                   
               
               
                 GDASDDEEPDEIEMLIRRLHTDILNDARNDPVEQAKKIRQARESQSDQTNSTTQLSVYERMILGSASQQSTDHQPG 
               
               
                   
               
               
                 EFSNMFRTLEDEQIEINQNFLFDEYDSEDDSIADDKVEIASDDEQMLLQEHKKRGKKYLQDEIVKEEDFDEDDDSDE 
               
               
                   
               
               
                 DIHMDDLENESLSFDRNNRKSHKPVCKRTREENILDADLGDEKDDEDTIFIDNLPSDEFSIRRQLQDVKSYIKQFEML 
               
               
                   
               
               
                 FFEEEDSDKEEQLKQITNVQKHEEALQNFKDRSHLKNFWCIPLSSDVREIDWDVLIARQQEHTNGQLFDVITCDPP 
               
               
                   
               
               
                 WQLSSANPTRGVAIAYETLNDGEILKIPWGRLQKDGFLFIWVINAKYRFALDMMGAHGYRVVDEIQWVKQTCNGKI 
               
               
                   
               
               
                 AKGHGYYLQHAKEVCLVGCKGDPAILAKKCRSNIESDVIFSERRGQSQKPEEIYELVEALVPNGYYMEIFGRRNNLH 
               
               
                   
               
               
                 NGWVTVGNEL 
               
               
                   
               
               
                 &gt;EJY79437.1 MT-A70 family protein [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 24) 
               
               
                 MHLPMQIITQNMFRQGNQHSCLNRTEILRTPRLTRSTKTELQEQTHFSKLPRRNYLKLQIDMREIQSLVDKKVKESA 
               
               
                   
               
               
                 AAQQQLSQSGIEDSAIKRSLRPRKVENYKNMLEGDEITLKTIQDEQIEVKRKKREASSQNRLEDEDEDEDMLEVGQ 
               
               
                   
               
               
                 QIERASDDEDDDDFPISTRRSARKRTRRQDVDEDEEAIEVNQVESSDAEVEIPANDIDTESYTEGTNKRKQKLKAKK 
               
               
                   
               
               
                 QVLDKKKNKTEGDIDKEDAVEEEETVFIDNLPNDEFEIRRMLKEVKKHIKSLEKQFFEEEDSEKEEELKQINNNSKHE 
               
               
                   
               
               
                 EALQAFKETSHLKQFWCIPLSVNVTTLDFDLLAKSQMKQGGRLFDVITIDPPWQLSSANPTRGVAIAYDTLNDKEILN 
               
               
                   
               
               
                 MPFEKVQTDGFLFIWVINAKYRFALEMMEKFGYKLVDEIAWVKQTVNGKIAKGHGYYLQHAKETCLVGVKGNVKGK 
               
               
                   
               
               
                 ARYNIESDVIFSQRRGQSQKPEEIYEIAEALVPNGYYLEIFGRRNNLHNGWVTIGNEL 
               
               
                   
               
               
                 &gt;NP_066012.1 N6-adenosine-methyltransferase non-catalytic subunit [ Homo sapiens ] 
               
               
                 (SEQ ID No: 25) 
               
               
                 MDSRLQEIRERQKLRRQLLAQQLGAESADSIGAVLNSKDEQREIAETRETCRASYDTSAPNAKRKYLDEGETDEDK 
               
               
                   
               
               
                 MEEYKDELEMQQDEENLPYEEEIYKDSSTFLKGTQSLNPHNDYCQHFVDTGHRPQNFIRDVGLADRFEEYPKLREL 
               
               
                   
               
               
                 IRLKDELIAKSNTPPMYLQADIEAFDIRELTPKFDVILLEPPLEEYYRETGITANEKCWTWDDIMKLEIDEIAAPRSFIFL 
               
               
                   
               
               
                 WCGSGEGLDLGRVCLRKWGYRRCEDICWIKTNKNNPGKTKTLDPKAVFQRTKEHCLMGIKGTVKRSTDGDFIHA 
               
               
                   
               
               
                 NVDIDLIITEEPEIGNIEKPVEIFHIIEHFCLGRRRLHLFGRDSTIRPGWLTVGPTLTNSNYNAETYASYFSAPNSYLTG 
               
               
                   
               
               
                 CTEEIERLRPKSPPPKSKSDRGGGAPRGGGRGGTSAGRGRERNRSNFRGERGGFRGGRGGAHRGGFPPR 
               
               
                   
               
               
                 &gt;NP_964000.2 N6-adenosine-methyltransferase non-catalytic [ Mus musculus ] 
               
               
                 (SEQ ID No: 26) 
               
               
                 MDSRLQEIRERQKLRRQLLAQQLGAESADSIGAVLNSKDEQREIAETRETCRASYDTSAPNSKRKCLDEGETDEDK 
               
               
                   
               
               
                 VEEYKDELEMQQEEENLPYEEEIYKDSSTFLKGTQSLNPHNDYCQHFVDTGHRPQNFIRDVGLADRFEEYPKLRELI 
               
               
                   
               
               
                 RLKDELIAKSNTPPMYLQADIEAFDIRELTPKFDVILLEPPLEEYYRETGITANEKCWTWDDIMKLEIDEIAAPRSFIFL 
               
               
                   
               
               
                 WCGSGEGLDLGRVCLRKWGYRRCEDICWIKTNKNNPGKTKTLDPKAVFQRTKEHCLMGIKGTVKRSTDGDFIHA 
               
               
                   
               
               
                 NVDIDLIITEEPEIGNIEKPVEIFHIIEHFCLGRRRLHLFGRDSTIRPGWLTVGPTLTNSNYNAETYASYFSAPNSYLTG 
               
               
                   
               
               
                 CTEEIERLRPKSPPPKSKSDRGGGAPRGGGRGGTSAGRGRERNRSNFRGERGGFRGGRGGTHRGGFTPR 
               
               
                   
               
               
                 &gt;XP_003129279.3 N6-adenosine-methyltransferase subunit METTL14 [ Sus scrofa ] 
               
               
                 (SEQ ID No: 27) 
               
               
                 MDSRLQEIRERQKLRRQLLAQQLGAESADSIGAVLNSKDEQREIAETRETCRASYDTSTPNAKRKYQDEGETDEDK 
               
               
                   
               
               
                 IEEYKDELEMQQEEENLPYEEEIYKDSSTFLKGTQSLNPHNDYCQHFVDTGHRPQNFIRDVGLADRFEEYPKLRELI 
               
               
                   
               
               
                 RLKDELIAKSNTPPMYLQADIEAFDIRELTPKFDVILLEPPLEEYYRETGITANEKCWTWDDIMKLEIDEIAAPRSFIFL 
               
               
                   
               
               
                 WCGSGEGLDLGRVCLRKWGYRRCEDICWIKTNKNNPGKTKTLDPKAVFQRTKEHCLMGIKGTVKRSTDGDFIHA 
               
               
                   
               
               
                 NVDIDLIITEEPEIGNIEKPVEIFHIIEHFCLGRRRLHLFGRDSTIRPGWLTVGPTLTNSNYNAETYASYFSAPNSYLTG 
               
               
                   
               
               
                 CTEEIERLRPKSPPPKSKSDRGGGAPRGGGRGGTSAGRGRERNRSNFRGERGGFRGGRGGAHRGGFPPR 
               
               
                   
               
               
                 &gt;XP_018099063.1 PREDICTED: N6-adenosine-methyltransferase subunit METTL14 
               
               
                 isoform X2 [ Xenopus laevis ] 
               
               
                 (SEQ ID No: 28) 
               
               
                 MNSRLQEIRARQTLRRKLLAQQLGAESADSIGAVLNSKDEQREIAETRETSRASYDTSAAVSKRKLPEEGKADEEV 
               
               
                   
               
               
                 VQECKDSVEPQKEEENLPYREEIYKDSSTFLKGTQSLNPHNDYCQHFVDTGHRPQNFIRDVGLADRFEEYPKLREL 
               
               
                   
               
               
                 IRLKDELIAKSNTPPMYLQADLENFDLRELKSEFDVILLEPPLEEYFRETGIAANEKWWTWEDIMKLDIEGIAGSRAFV 
               
               
                   
               
               
                 FLWCGSGEGLDFGRMCLRKWGFRRSEDICWIKTNKDNPGKTKTLDPKAIFQRTKEHCLMGIKGTVHRSTDGDFIH 
               
               
                   
               
               
                 ANVDIDLIITEEPEIGNIEKPVEIFHIIEHFCLGRRRLHLFGRDSTIRPDQSWEERLANSGGLREKEFLVGLLLGLLLPTA 
               
               
                   
               
               
                 TLIQRLMLLTLTLQIHLLLDAQRRSKDSVPKLHLLSQIVALGHREEEDEVEHLQVAERGAGKGTEAVLGETEGISEDV 
               
               
                   
               
               
                 EDHIGVSLLPVDFKCF 
               
               
                   
               
               
                 &gt;NP_996954.1 N6-adenosine-methyltransferase non-catalytic subunit [ Danio rerio ] 
               
               
                 (SEQ ID No: 29) 
               
               
                 MNSRLQEIRERQKLRRQLLAQQLGAESPDSIGAVLNSKDEQKEIEETRETCRASFDISVPGAKRKCLNEGEDPEED 
               
               
                   
               
               
                 VEEQKEDVEPQHQEESGPYEEVYKDSSTFLKGTQSLNPHNDYCQHFVDTGHRPQNFIRDGGLADRFEEYPKQRE 
               
               
                   
               
               
                 LIRLKDELISATNTPPMYLQADPDTFDLRELKCKFDVILIEPPLEEYYRESGIIANERFWNWDDIMKLNIEEISSIRSFVF 
               
               
                   
               
               
                 LWCGSGEGLDLGRMCLRKWGFRRCEDICWIKTNKNNPGKTKTLDPKAVFQRTKEHCLMGIKGTVRRSTDGDFIH 
               
               
                   
               
               
                 ANVDIDLIITEEPEMGNIEKPVEIFHIIEHFCLGRRRLHLFGRDSTIRPGWLTVGPTLTNSNFNIEVYSTHFSEPNSYLS 
               
               
                   
               
               
                 GCTEEIERLRPKSPPPKSMAERGGGAPRGGRGGPAAGRGDRGRERNRPNFRGDRGGFRGRGGPHRGFPPR 
               
               
                   
               
               
                 &gt;NP_609205.1 methyltransferase like 14 [ Drosophila melanogaster ] 
               
               
                 (SEQ ID No: 30) 
               
               
                 MSDVLKSSQERSRKRRLLLAQTLGLSSVDDLKKALGNAEDINSSRQLNSGGQREEEDGGASSSKKTPNEIIYRDSS 
               
               
                   
               
               
                 TFLKGTQSSNPHNDYCQHFVDTGQRPQNFIRDVGLADRFEEYPKLRELIKLKDKLIQDTASAPMYLKADLKSLDVKT 
               
               
                   
               
               
                 LGAKFDVILIEPPLEEYARAAPSVATVGGAPRVFWNWDDILNLDVGEIAAHRSFVFLWCGSSEGLDMGRNCLKKW 
               
               
                   
               
               
                 GFRRCEDICWIRTNINKPGHSKQLEPKAVFQRTKEHCLMGIKGTVRRSTDGDFIHANVDIDLIISEEEEFGSFEKPIEI 
               
               
                   
               
               
                 FHIIEHFCLGRRRLHLFGRDSSIRPGWLTVGPELTNSNFNSELYQTYFAEAPATGCTSRIELLRPKSPPPNSKVLRG 
               
               
                   
               
               
                 RGRGFPRGRGRPR 
               
               
                   
               
               
                 &gt;NP_567348.2 Methyltransferase MT-A70 family protein [ Arabidopsis thaliana ] 
               
               
                 (SEQ ID No: 31) 
               
               
                 MKKKQEESSLEKLSTWYQDGEQDGGDRSEKRRMSLKASDFESSSRSGGSKSKEDNKSVVDVEHQDRDSKRERD 
               
               
                   
               
               
                 GRERTHGSSSDSSKRKRWDEAGGLVNDGDHKSSKLSDSRHDSGGERVSVSNEHGESRRDLKSDRSLKTSSRDE 
               
               
                   
               
               
                 KSKSRGVKDDDRGSPLKKTSGKDGSEVVREVGRSNRSKTPDADYEKEKYSRKDERSRGRDDGWSDRDRDQEGL 
               
               
                   
               
               
                 KDNWKRRHSSSGDKDQKDGDLLYDRGREREFPRQGRERSEGERSHGRLGGRKDGNRGEAVKALSSGGVSNEN 
               
               
                   
               
               
                 YDVIEIQTKPHDYVRGESGPNFARMTESGQQPPKKPSNNEEEWAHNQEGRQRSETFGFGSYGEDSRDEAGEASS 
               
               
                   
               
               
                 DYSGAKARNQRGSTPGRTNFVQTPNRGYQTPQGTRGNRPLRGGKGRPAGGRENQQGAIPMPIMGSPFANLGMP 
               
               
                   
               
               
                 PPSPIHSLTPGMSPIPGTSVTPVFMPPFAPTLIWPGARGVDGNMLPVPPVLSPLPPGPSGPRFPSIGTPPNPNMFFT 
               
               
                   
               
               
                 PPGSDRGGPPNFPGSNISGQMGRGMPSDKTSGGWVPPRGGGPPGKAPSRGEQNDYSQNFVDTGMRPQNFIRE 
               
               
                   
               
               
                 LELTNVEDYPKLRELIQKKDEIVSNSASAPMYLKGDLHEVELSPELFGTKFDVILVDPPWEEYVHRAPGVSDSMEYW 
               
               
                   
               
               
                 TFEDIINLKIEAIADTPSFLFLWVGDGVGLEQGRQCLKKWGFRRCEDICWVKTNKSNAAPTLRHDSRTVFQRSKEH 
               
               
                   
               
               
                 CLMGIKGTVRRSTDGHIIHANIDTDVIIAEEPPYGSTQKPEDMYRIIEHFALGRRRLELFGEDHNIRAGWLTVGKGLSS 
               
               
                   
               
               
                 SNFEPQAYVRNFADKEGKVWLGGGGRNPPPDAPHLVVTTPDIESLRPKSPMKNQQQQSYPSSLASANSSNRRTT 
               
               
                   
               
               
                 GNSPQANPNVVVLHQEASGSNFSVPTTPHWVPPTAPAAAGPPPMDSFRVPEGGNNTRPPDDKSFDMYGFN 
               
               
                   
               
               
                 &gt;PNW88915.1 hypothetical protein CHLRE_01g050600v5 [ Chlamydomonas reinhardtii ] 
               
               
                 (SEQ ID No: 32) 
               
               
                 MQDGQGPPGDGRGRGRGRSRGGRIMFAREGGRGPRPMHSDMGPPPPPMGMFPHDPSAMMGGPMPGMPPM 
               
               
                   
               
               
                 DFTPEMLLTMMGAGLGGPMGLAGPMGMMMPDFGAAAAGAPGGMMVPPGAMMPPPPQPPSGGPGGMGGGGM 
               
               
                   
               
               
                 GGMGGMMGHQQGMGGAGGPMGLPGGGMGMGMGGGGGGGGGGGYGGRGGHGEAGGGGGGGGRAGGAG 
               
               
                   
               
               
                 GGGGAGGAAEHLSNDYSQNFVDTGLRPQNFLRDTHLTDRYEEYPKLKELIVRKDRQVSAHATPPLFLRTDLRSTRL 
               
               
                   
               
               
                 SPELFGTKFDVILVDPPWEEYVRRAPGMVADPEVWSWQDIQALDIEAVADNPCFLFLWCGAEEGLEAGRVCMQK 
               
               
                   
               
               
                 WGFRRVEDICWIKTNKEGGKGPGGGRRPYLTAANQHPESMLVHTKEHCLMGIKGSVRRATDGHIIHTNVDTDVIV 
               
               
                   
               
               
                 SEEPELGSTRKPEEMYHIIERFCNGRRRLELFGEDHNIRNGWVTVGRSLTSSNFSAKAYADHFRNRDGSVWVQNT 
               
               
                   
               
               
                 YGPKPPPGSVILVPTTDEIEDLRPKSPTGPHGGSSFHHSR 
               
               
                   
               
               
                 &gt;XP_001022374.1 MT-a70 family protein [ Tetrahymena thermophila  SB210] 
               
               
                 (SEQ ID No: 33) 
               
               
                 MQPQQNQNQQQQQQQQSQQQQQQNQQLPQLQQSMSSQQQQNQQQEKQIIIKRGTTSKRNDYCQNFVNTHER 
               
               
                   
               
               
                 PQNFIMNIRPEERFIEYPKLQDLIKFKDDLIKKRNHPPVYLKADLKYYDLSKLGKFDVIMMDPPWKEYEERVQGLPIYS 
               
               
                   
               
               
                 QYPEKFNSWDLNEIAALPIDEISDKPSFLFLWVGSDHLDQGRELFRKWGYKRCEDIVWVKTNKDKTKEYIELPHSNL 
               
               
                   
               
               
                 LVRVKEHCLVGLRGDVKRASDSHFIHANIDTDVIVAEEPPLGSTQKPAEIYDIIERFCLGRKRLELFGEVHNVRQGWL 
               
               
                   
               
               
                 TIGKLLDESNFNQDEYNSWFDGDKTYPQIQTYRGGRYVGTTPDIEQLRPKSPTKNNQMNSNQNMSGSQVSEFDL 
               
               
                   
               
               
                 GIQQKQQKLNQQF 
               
               
                   
               
               
                 &gt;NP_009876.1 Kar4p [ Saccharomyces cerevisiae  S288C] 
               
               
                 (SEQ ID No: 34) 
               
               
                 MAFQDPTYDQNKSRHINNSHLQGPNQETIEMKSKHVSFKPSRDFHTNDYSNNYIHGKSLPQQHVTNIENRVDGYP 
               
               
                   
               
               
                 KLQKLFQAKAKQINQFATTPFGCKIGIDSIVPTLNHWIQNENLTFDVVMIGCLTENQFIYPILTQLPLDRLISKPGFLFI 
               
               
                   
               
               
                 WANSQKINELTKLLNNEIWAKKFRRSEELVFVPIDKKSPFYPGLDQDDETLMEKMQWHCWMCITGTVRRSTDGHLI 
               
               
                   
               
               
                 HCNVDTDLSIETKDTTNGAVPSHLYRIAENFSTATRRLHIIPARTGYETPVKVRPGWVIVSPDVMLDNFSPKRYKEEI 
               
               
                   
               
               
                 ANLGSNIPLKNEIELLRPRSPVQKAQ 
               
               
                   
               
               
                 &gt;XP_001691478.1 predicted protein [ Chlamydomonas reinhardtii ] 
               
               
                 (SEQ ID No: 35) 
               
               
                 MRLGGGPGGSELDDLLGKRSVKEKVKVEKGSELLDILSKPTARESARVEQFRTAGGSAIREHCPHLTKDECRRVN 
               
               
                   
               
               
                 GVPLACHRLHFLRVVQPHTDVALGNCSYLDTCRNMRTCKYVHYRPDPEPDVPGMGSEMARLRASVPKKPVGDG 
               
               
                   
               
               
                 QTSRGALDPQWINCDVRSFDMTVLGKFGVIMADPPWEIHQDLPYGTMKDDEMVNLNVGCLQDNGVLFLWVTGRA 
               
               
                   
               
               
                 MELARECMAKWGYKRVDELIWVKTNQLQRLIRTGRTGHWLNHSKEHCLVGIKGSPQLNRYVDTDVVVAEVRETS 
               
               
                   
               
               
                 RKPDEMYSLLERLSPGTRKLEIFARVHNCKPGWVGLGNQLKNVNLIEPEVRQRFAARYGFEPDASKDCFVN 
               
               
                   
               
               
                 &gt;NP_192814.1 mRNAadenosine methylase [ Arabidopsis thaliana ] 
               
               
                 (SEQ ID No: 36) 
               
               
                 METESDDATITVVKDMRVRLENRIRTQHDAHLDLLSSLQSIVPDIVPSLDLSLKLISSFTNRPFVATPPLPEPKVEKKH 
               
               
                   
               
               
                 HPIVKLGTQLQQLHGHDSKSMLVDSNQRDAEADGSSGSPMALVRAMVAECLLQRVPFSPTDSSTVLRKLENDQNA 
               
               
                   
               
               
                 RPAEKAALRDLGGECGPILAVETALKSMAEENGSVELEEFEVSGKPRIMVLAIDRTRLLKELPESFQGNNESNRVVE 
               
               
                   
               
               
                 TPNSIENATVSGGGFGVSGSGNFPRPEMWGGDPNMGFRPMMNAPRGMQMMGMHHPMGIMGRPPPFPLPLPLP 
               
               
                   
               
               
                 VPSNQKLRSEEEDLKDVEALLSKKSFKEKQQSRTGEELLDLIHRPTAKEAATAAKFKSKGGSQVKYYCRYLTKEDC 
               
               
                   
               
               
                 RLQSGSHIACNKRHFRRLIASHTDVSLGDCSFLDTCRHMKTCKYVHYELDMADAMMAGPDKALKPLRADYCSEAE 
               
               
                   
               
               
                 LGEAQWINCDIRSFRMDILGTFGVVMADPPWDIHMELPYGTMADDEMRTLNVPSLQTDGLIFLWVTGRAMELGRE 
               
               
                   
               
               
                 CLELWGYKRVEEIIWVKTNQLQRIIRTGRTGHWLNHSKEHCLVGIKGNPEVNRNIDTDVIVAEVRETSRKPDEMYA 
               
               
                   
               
               
                 MLERIMPRARKLELFARMHNAHAGWLSLGNQLNGVRLINEGLRARFKASYPEIDVQPPSPPRASAMETDNEPMAID 
               
               
                   
               
               
                 SITA 
               
               
                   
               
               
                 &gt;EAS00013.2 N6-adenosine-methyltransferase 70 kDa subunit [ Tetrahymena thermophila  SB210] 
               
               
                 (SEQ ID No: 37) 
               
               
                 MGSSVKDQEISNKKHKARNSSSGANNNSNSSNYQSSKRDIHQDRSYSKDDSQSRQYNSNNGGGGSSSKNSNRN 
               
               
                   
               
               
                 SSQQGYNQNSSSNQGQNSEYGGSGSGKNSQANSQRNSSQQGLQQLNQQQQSQQQQQQMLQNQMNSMGMM 
               
               
                   
               
               
                 NQFQNSFGLMGMQPSQPLQLLNPSMIIPSGKKQKYDFLEFPPSSQHEFRAILLDYFLSDLFDYPMHSAELFENFIEA 
               
               
                   
               
               
                 FSDIKDSSSFIKKLELIPLLQELNDKKAIKLETCAVGTKLFDFIVDINKDKIKQLSREFSKDRPKFMPILDKKPQPSSSKT 
               
               
                   
               
               
                 NSSSTTAPPKQAISKREIEDLLKKETGLQKEVITQSKEKSNLLNKISAAEESALAIFRKQGSRRIDYCDCGTRDKCIQIR 
               
               
                   
               
               
                 NSTVPCNKAHFRKIIRPHTDENLGNCSYLDTCRHMDYCKFVHYELDVDINNMNNDNLLLDGIEKKLNPQWINCDLR 
               
               
                   
               
               
                 QIDFNILGKFNCIMADPPWDIHMTLPYGTLKDREMKAMRVDLLQEEGVIFLWVTGRAMELGRECLTNWGYRRVEEI 
               
               
                   
               
               
                 IWVKTNQLQRIIRTGRTGHWLNHSKEHCLVGIKGNPKINRKIDCDVIVSEVRETSRKPDEIYNLIERMCPGGKKIELFG 
               
               
                   
               
               
                 RPHNTMPGWLTLGNQLPGIYLEDEEIIERYMDAYPDQDISRETMERNRIRMKNENDIDHIYNSHIQNIPPFKTKQLTK 
               
               
                   
               
               
                 DLQLQQQSSSMQTTQQQSSSQMMPQMQQQQSSQSINSNTDLQMHGNGLYEQE 
               
               
                   
               
               
                 &gt;ORX92345.1 MT-A70-domain-containing protein [ Basidiobolus meristosporus  CBS 931.73] 
               
               
                 (SEQ ID No: 38) 
               
               
                 MKLERALFKMADMWGYNTIGIKREYDNDKSAISVIYFDPRNLRNVQHIEKTLEDICDVDSIDPDIFLDKTTSAQVPSTY 
               
               
                   
               
               
                 IPNEEARFSEDAEIEKLLSKPSFLEMEAFSSLIGVTELIERKTFREQEAEEMFKAQGNGGFREFCEYLIKEDCKKMNT 
               
               
                   
               
               
                 SGQPCAMTASILLTNMKLHFRRIMRPQTDLELGDCSYLNTCHRMDTCKYVHYELDDFEHPSSANITKTTIPTSLIFRP 
               
               
                   
               
               
                 PKKVLPAQWINCDVRKFDFSILGKFSVIMADPPWDIHMTLPYGTMTDDEMKAMAIHKLQDEGLIFLWVTARAMELG 
               
               
                   
               
               
                 RECLATWGYDRVDEVVWIKTNQLQRLIRTGRTGHWLNHSKEHCLVGIKGDPSRFNIGLACDVLVAEVRETSRKPD 
               
               
                   
               
               
                 QIYGMIDRLSPGTRKIEIFGRQHNTRPGWFTLGNQLKDVRIVEPEVLEAYNQRYPECPAQLSAIPES 
               
               
                   
               
               
                 &gt;AJR96662.1 Ime4p [ Saccharomyces cerevisiae  YJM1248] 
               
               
                 (SEQ ID No: 39) 
               
               
                 MINDKLVHFLIQNYDDILRAPLSGQLKDVYSLYISGGYDDEMQKLRNDKDEVLQFEQFWNDLQDIIFATPQSIQFDQN 
               
               
                   
               
               
                 LLVADRPEKIVYLDVFSLKILYNKFHAFYYTLKSSSSSCEEKVSSLTTKPEADSEKDQLLGRLLGVLNWDVNVSNQGL 
               
               
                   
               
               
                 PREQLSNRLQNLLREKPSSFQLAKERAKYTTEVIEYIPICSDYSHASLLSTAVYIVNNKIVSLQWSKISACQENHPGLI 
               
               
                   
               
               
                 ECIQSKIHFIPNIKPQTDISLGDCSYLDTCHKLNMCRYIHYLQYIPSCLQERADRETAIENKRIRSNVSIPFYTLGNCSA 
               
               
                   
               
               
                 HCIKKALPAQWIRCDVRKFDFRVLGKFSVVIADPAWNIHMNLPYGTCNDIELLGLPLHELQDEGIIFLWVTGRAIELG 
               
               
                   
               
               
                 KESLNNWGYNVINEVSWIKTNQLGRTIVTGRTGHWLNHSKEHLLVGLKGNPKWINKHIDVDLIVSMTRETSRKPDE 
               
               
                   
               
               
                 LYGIAERLAGTHARKLEIFGRDHNTRPGWFTIGNQLTGNCIYEMDVERKYQEFMKSKTGTSHTGTKKIDKKQPSKL 
               
               
                   
               
               
                 QQQHQQQYWNNMDMGSGKYYAEAKQNPMNQKHTPFESKQQQKQQFQTLNNLYFAQ 
               
               
                   
               
               
                 &gt;NP_651204.1 methyltransferase like 3 [ Drosophila melanogaster ] 
               
               
                 (SEQ ID No: 40) 
               
               
                 MADAWDIKSLKTKRNTLREKLEKRKKERIEILSDIQEDLTNPKKELVEADLEVQKEVLQALSSCSLALPIVSTQVVEKI 
               
               
                   
               
               
                 AGSSLEMVNFILGKLANQGAIVIRNVTIGTEAGCEIISVQPKELKEILEDTNDTCQQKEEEAKRKLEVDDVDQPQEKTI 
               
               
                   
               
               
                 KLESTVARKESTSLDAPDDIMMLLSMPSTREKQSKQVGEEILELLTKPTAKERSVAEKFKSHGGAQVMEFCSHGTK 
               
               
                   
               
               
                 VECLKAQQATAEMAAKKKQERRDEKELRPDVDAGENVTGKVPKTESAAEDGEIIAEVINNCEAESQESTDGSDTCS 
               
               
                   
               
               
                 SETTDKCTKLHFKKIIQAHTDESLGDCSFLNTCFHMATCKYVHYEVDTLPHINTNKPTDVKTKLSLKRSVDSSCTLYP 
               
               
                   
               
               
                 PQWIQCDLRFLDMTVLGKFAVVMADPPWDIHMELPYGTMSDDEMRALGVPALQDDGLIFLWVTGRAMELGRDCL 
               
               
                   
               
               
                 KLWGYERVDELIWVKTNQLQRIIRTGRTGHWLNHGKEHCLVGMKGNPTNLNRGLDCDVIVAEVRATSHKPDEIYGI 
               
               
                   
               
               
                 IERLSPGTRKIELFGRPHNIQPNWITLGNQLDGIRLVDPELITQFQKRYPDGNCMSPASANAASINGIQK 
               
               
                   
               
               
                 &gt;NP_001084701.1 methyltransferase like 3 L homeolog [ Xenopus laevis ] 
               
               
                 (SEQ ID No: 41) 
               
               
                 MSDTWSSIQAHKKQLDNLRERLQRRRKDATSQLALDLQSSEGGIAPTFRSDSPVPSASSQPLKGPSGSAEVTPDP 
               
               
                   
               
               
                 ELEKKLLHHLSDLSLVLPADSVSIQLAITTPDFPVTRQGVESLLQKFAAQELIEVKGWGQEDDDRPTVVTFADYSKLS 
               
               
                   
               
               
                 AMMGAVAERKGTTIPTGAKKRRLQEADPSASSLSSSLSASASREKKTSEPQKKARKHASHLDLEIESLLSQQSTKE 
               
               
                   
               
               
                 QQSKKVSQEILELLSTSTAKEQSIVEKFRSRGRAQVQEFCDFGTKEECMKAAGADTPCRKLHFRRIINMHTDESLG 
               
               
                   
               
               
                 DCSFLNTCFHMDTCKYVHYEIDAWVEPGGTAMGTEAIASLDTPLAKAVGDSSVGRLFPAQWIRCDIRYLDVSILGKF 
               
               
                   
               
               
                 SVVMADPPWDIHMELPYGTLTDDEMRKLQIPVLQDDGFLFLWVTGRAMELGRECLKLWGYERVDEIIWVKTNQLQ 
               
               
                   
               
               
                 RIIRTGRTGHWLNHGKEHCLVGVKGSPQGFNRGLDCDVIVAEVRSTSHKPDEIYGMIERLSPGTRKIELFGRPHNIQ 
               
               
                   
               
               
                 PNWITLGNQLDGIHLLDPDVVAQFKQKYPDGVIGMPKNM 
               
               
                   
               
               
                 sp|F1R777.1|MTA70_DANRE RecName: Full = N6-adenosine-methyltransferase subunit METTL3: 
               
               
                 AltName: Full = N6-adenosine-methyltransferase 70 kDa subunit; Short = MT-A70  
               
               
                 (SEQ ID No: 416) 
               
               
                 MSDTWSHIQAHKKQLDSLRERLQRRRKDPTQLGTEVGSVESGSARSDSPGPAIQSPPQVEVEHPPDPELEKRLLG 
               
               
                   
               
               
                 YLSELSLSLPTDSLTITNQLNTSESPVSHSCIQSLLLKFSAQELIEVRQPSITSSSSSTLVTSVDHTKLWAMIGSAGQS 
               
               
                   
               
               
                 QRTAVKRKADDITHQKRALGSSPSIQAPPSPPRKSSVSLATASISQLTASSGGGGGGADKKGRSNKVQASHLDMEI 
               
               
                   
               
               
                 ESLLSQQSTKEQQSKKVSQEILELLNTSSAKEQSIVEKFRSRGRAQVQEFCDYGTKEECVQSGDTPQPCTKLHFRR 
               
               
                   
               
               
                 IINKHTDESLGDCSFLNTCFHMDTCKYVHYEIDSPPEAEGDALGPQAGAAELGLHSTVGDSNVGKLFPSQWICCDIR 
               
               
                   
               
               
                 YLDVSILGKFAVVMADPPWDIHMELPYGTLTDDEMRKLNIPILQDDGFLFLWVTGRAMELGRECLSLWGYDRVDEII 
               
               
                   
               
               
                 WVKTNQLQRIIRTGRTGHWLNHGKEHCLVGVKGNPQGFNRGLDCDVIVAEVRSTSHKPDEIYGMIERLSPGTRKIE 
               
               
                   
               
               
                 LFGRPHNVQPNWITLGNQLDGIHLLDPEVVARFKKRYPDGVISKPKNM 
               
               
                   
               
               
                 &gt;NP_062826.2 N6-adenosine-methyltransferase catalytic subunit [ Homo sapiens ] 
               
               
                 (SEQ ID No: 42) 
               
               
                 MSDTWSSIQAHKKQLDSLRERLQRRRKQDSGHLDLRNPEAALSPTFRSDSPVPTAPTSGGPKPSTASAVPELATD 
               
               
                   
               
               
                 PELEKKLLHHLSDLALTLPTDAVSICLAISTPDAPATQDGVESLLQKFAAQELIEVKRGLLQDDAHPTLVTYADHSKLS 
               
               
                   
               
               
                 AMMGAVAEKKGPGEVAGTVTGQKRRAEQDSTTVAAFASSLVSGLNSSASEPAKEPAKKSRKHAASDVDLEIESLL 
               
               
                   
               
               
                 NQQSTKEQQSKKVSQEILELLNTTTAKEQSIVEKFRSRGRAQVQEFCDYGTKEECMKASDADRPCRKLHFRRIINK 
               
               
                   
               
               
                 HTDESLGDCSFLNTCFHMDTCKYVHYEIDACMDSEAPGSKDHTPSQELALTQSVGGDSSADRLFPPQWICCDRY 
               
               
                   
               
               
                 LDVSILGKFAVVMADPPWDIHMELPYGTLTDDEMRRLNIPVLQDDGFLFLWVTGRAMELGRECLNLWGYERVDEII 
               
               
                   
               
               
                 WVKTNQLQRIIRTGRTGHWLNHGKEHCLVGVKGNPQGFNQGLDCDVIVAEVRSTSHKPDEIYGMIERLSPGTRKIE 
               
               
                   
               
               
                 LFGRPHNVQPNWITLGNQLDGIHLLDPDVVARFKQRYPDGIISKPKNL 
               
               
                   
               
               
                 &gt;sp|Q8C3P7.2|MTA70_MOUSE RecName: Full = N6-adenosine-methyltransferase subunit METTL3; 
               
               
                 AltName: Full = Methyltransferase-like protein 3; AltName: Full = N6-adenosine- 
               
               
                 methyltransferase 70 kDa subunit; Short = MT-A70 
               
               
                 (SEQ ID No: 43) 
               
               
                 MSDTWSSIQAHKKQLDSLRERLQRRRKQDSGHLDLRNPEAALSPTFRSDSPVPTAPTSSGPKPSTTSVAPELATD 
               
               
                   
               
               
                 PELEKKLLHHLSDLALTLPTDAVSIRLAISTPDAPATQDGVESLLQKFAAQELIEVKRGLLQDDAHPTLVTYADHSKLS 
               
               
                   
               
               
                 AMMGAVADKKGLGEVAGTIAGQKRRAEQDLTTVTTFASSLASGLASSASEPAKEPAKKSRKHAASDVDLEIESLLN 
               
               
                   
               
               
                 QQSTKEQQSKKVSQEILELLNTTTAKEQSIVEKFRSRGRAQVQEFCDYGTKEECMKASDADRPCRKLHFRRIINKH 
               
               
                   
               
               
                 TDESLGDCSFLNTCFHMDTCKYVHYEIDACVDSESPGSKEHMPSQELALTQSVGGDSSADRLFPPQWICCDIRYL 
               
               
                   
               
               
                 DVSILGKFAVVMADPPWDIHMELPYGTLTDDEMRRLNIPVLQDDGFLFLWVTGRAMELGRECLNLWGYERVDEIIW 
               
               
                   
               
               
                 VKTNQLQRIIRTGRTGHWLNHGKEHCLVGVKGNPQGFNQGLDCDVIVAEVRSTSHKPDEIYGMIERLSPGTRKIEL 
               
               
                   
               
               
                 FGRPHNVQPNWITLGNQLDGIHLLDPDVVARFKQRYPDGIISKPKNL 
               
               
                   
               
               
                 &gt;XP_003128628.1 N6-adenosine-methyltransferase 70 kDa subunit [ Sus scrofa ] 
               
               
                 (SEQ ID No: 44) 
               
               
                 MSDTTWSSIQAHKKQLDSLRERLRRRRKQDSGHLDLRNPEAALSPTFRSDSPVPTVPTSGGPKPSTASAVPELATD 
               
               
                   
               
               
                 PELEKKLLHHLSDLALTLPTDAVSIRLAISTPDAPATQDGVESLLQKFAAQELIEVKRSLLQDDAHPTLVTYADHSKLS 
               
               
                   
               
               
                 AMMGAVAEKKGPGEVAGTITGQKRRAEQDSTTVAAFASSLTSSLASSASEVAKEPTKKSRKHAASDVDLEIESLLN 
               
               
                   
               
               
                 QQSTKEQQSKKVSQEILELLNTTTAKEQSIVEKFRSRGRAQVQEFCDYGTKEECMKASDADRPCRKLHFRRIINKH 
               
               
                   
               
               
                 TDESLGDCSFLNTCFHMDTCKYVHYEIDACMDSEAPGSKDHTPSQELALTQSVGGDSNADRLFPPQWICCDIRYL 
               
               
                   
               
               
                 DVSILGKFAVVMADPPWDIHMELPYGTLTDDEMRRLNIPVLQDDGFLFLWVTGRAMELGRECLNLWGYERVDEIIW 
               
               
                   
               
               
                 VKTNQLQRIIRTGRTGHWLNHGKEHCLVGVKGNPQGNQGLDCDVIVAEVRSTSHKPDEIYGMIERLSPGTRKIEL 
               
               
                   
               
               
                 FGRPHNVQPNWITLGNQLDGIHLLDPDVVARFKQRYPDGIISKPKNL 
               
               
                   
               
               
                 &gt;WP_009339935.1 MULTISPECIES: S-adenosylmethionine-binding protein [ Afipia ] 
               
               
                 (SEQ ID No: 45) 
               
               
                 MTLPAKDLLSFAGQRRFSTILADPPWQFTNKTGKVAPEHKRLSRYGTMKLDEIMMLPVADIAAPTSHLYLWCPNAL 
               
               
                   
               
               
                 LPEGLAVMKAWGFNYKSNIVWHKVRKDGGSDGRGVGFYFRNVTEVILFGVRGKNARTLAPGRRQVNLLATRKRE 
               
               
                   
               
               
                 HSRKPDEQYEIIESCSPGPFLELFARGTRKNWATWGNQADDDYKPTWKTYAHHSRAGLVAAE 
               
               
                   
               
               
                 &gt;WP_013485562.1 S-adenosylmethionine-binding protein [ Ethanoligenens harbinense ] 
               
               
                 (SEQ ID No: 46) 
               
               
                 MSTAKETANNLLQFCGEKKYATVYADPPWRFQNRTGKVAPENKKLNRYPTMDLEDIKALPVGKIAAEKSHLYLWVP 
               
               
                   
               
               
                 NALLPDGLEVMKAWGFEYKGNIIWEKVRKDGEPDGRGVGFYFRNVTEILLFGIRGGNNRTLAPARSQVNLIRTQKR 
               
               
                   
               
               
                 EHSRKPDEIITIIESCSPGPYLELFARGDRENWDMWGNQATAEYEPTWNTYKNHTTKETTSGVSGSQSET 
               
               
                   
               
               
                 &gt;WP_016343787.1 adenine-specific DNA methyltransferase [ Mycobacteroides abscessus ] 
               
               
                 (SEQ ID No: 47) 
               
               
                 MAAPLREVNEPPPLPVTDGGFSTILADPPWRFTNRTGKVAPEHRRLDRYSTLSLDEICALGVSDVTADNAHLYLWV 
               
               
                   
               
               
                 PNALLPDGLRVMEEWGFRYVSNIVWSKVRRDGLPDGRGVGFYFRNTTELLLFGVRGSMRTLQPARSQVNQIVTR 
               
               
                   
               
               
                 KREHSRKPDEQYELIEACSPGPYLEMFGRYRRPNWAVWGDEANEDVEPRGQTHKGYGGGEITRLPALEPHSRIP 
               
               
                   
               
               
                 QWLAKPIAAAIKSAYDDGMSIDAIAAETGYSISRVRHLLDQAGAKKRGRGRPAKA 
               
               
                   
               
               
                 &gt;WP_023133224.1 MULTISPECIES: MT-A70 protein [ Rothia ]  
               
               
                 (SEQ ID No: 48) 
               
               
                 MLDPMNTNEEFAPLPTVEGGFQTVLADPPWRFTNRTGKVAPEHHRLGRYGTMSLDEIKALRVGDVTADNAHLYL 
               
               
                   
               
               
                 WVPNALLPEGLEVMQAWGFRYVSNIIWAKRRKDGGPDGRGVGFYFRNVTEPILFGVKGSMRTLAPGRSTVNMIET 
               
               
                   
               
               
                 RKREHSRKPDEQYDLIEACSPGPYLELFARYARPGWSVWGNEASNEIEPRGKAQKGYGGGEIDRLPILEPNERMS 
               
               
                   
               
               
                 EWLSGRVGELLAEEYTKGASVQELANQSGYSIARVRTLLTHSGVPLRGRGRPKKGQVAS 
               
               
                   
               
               
                 &gt;ETW92643.1 S-adenosylmethionine-binding protein [ Candidatus Entotheonella  factor]  
               
               
                 (SEQ ID No: 49) 
               
               
                 MSNSPHSAADDLLACGFPPHSFSTVLADPPWRFTNRTGKMAPEHRRLSRYPTLTLEEIADLPLAQLVQPDSHLYLW 
               
               
                   
               
               
                 VPNALLAEGLDVMRRWGFTYKTNLVWYKIRRDGGPDRRGVGFYFRNVTELVLFGVRGRMRTLAPGRRQENLLAS 
               
               
                   
               
               
                 QKQEHSRKPDTFYDLIERCSPGPYLELFARHPRPGWHQFGNEPLVSSS 
               
               
                   
               
               
                 &gt;AHJ63281.1 Adenine-specific methyltransferase [ Granulibacter bethesdensis ] 
               
               
                 (SEQ ID No: 50) 
               
               
                 MTKQPDPIAEFRNQLNGGNFATVLADPPWRFQNRTGKMAPEHRRLSRYGTMELPEIMALPVSEVTAKTAHLYLWV 
               
               
                   
               
               
                 PNALLPEGLAVMQAWGFNYKSNLVWHKIRKDGGSDGRGVGFYFRNVTELVLFGVKGKNARTEAPGRRQVNLLAT 
               
               
                   
               
               
                 QKREHSRKPDEFYDIVEACSPGPYLELFARGTRPGWCAWGNQAEEYDITWDTYSHHSQRQSLWVAE 
               
               
                   
               
               
                 &gt;WP_017364718.1 S-adenosylmethionine-binding protein [ Methylococcus capsulatus ] 
               
               
                 (SEQ ID No: 51) 
               
               
                 MTENTLDPAADLLERLGDKRFRTILADPPWQFQNRTGKMAPEHKRLNRYGTMSLEAIAGLPVERLTADTAHLYLWV 
               
               
                   
               
               
                 PNALLLEGLKVMEAWGFTYKTNLVWHKIRKDGGPDGRGVGFYFRNVTELVLFGVRGKNARTLAAGRRQVNFLAT 
               
               
                   
               
               
                 RKREHSRKPDEMYGIIEACSPGPYLELFARGARDRWSVWGNEADENYYPRWNTYANHSQAEICPFE 
               
               
                   
               
               
                 &gt;WP_027700599.1 S-adenosylmethionine-binding protein [ Xylella fastidiosa ] 
               
               
                 (SEQ ID No: 52) 
               
               
                 MTKHKANTASDVGRDLLARHGGQRFHTILADPPWQFQNRTGKMAPEHKRLSRYGTMTLDDIMMLPVEQLVTDTA 
               
               
                   
               
               
                 HLYLWVPNALLPEGIKVLEAWGFSYKSNIVWHKVRKDGGPDGRGVGFYFRNVTELVLFGVRGKNARTLAPGRRQ 
               
               
                   
               
               
                 VNFLATQKREHSRKPDEFYDIVESCSPGPFLELFARGPRDGWKVWGNQADKYYPTWPTYSNHSQAECELGRVE 
               
               
                   
               
               
                 MIAQRLLSV 
               
               
                   
               
               
                 &gt;WP_027488351.1 S-adenosylmethionine-binding protein [ Rhizobium undicola ] 
               
               
                 (SEQ ID No: 53) 
               
               
                 MLNRNTDAPSPSDDFTNFISGRKFATIMADPPWQFMNRTGKVAPEHKRLNRYGTMELDAIKALPVATACAPTAHLY 
               
               
                   
               
               
                 LWVPNALLPEGLEVMKAWGFNYKANIVWHKLRKDGGSDGRGVGFYFRNVTELILFGTRGKNARTLPPGRSQVNYI 
               
               
                   
               
               
                 GTRKREHSRKPDEQYPLIESCSPGPYLEMFGRGLRKGWTTWGNQADETYEPTWKTYGHNSSTDRLEAAE 
               
               
                   
               
               
                 &gt;ESK34829.1 hypothetical protein G966_02949 [ Escherichia coli  UMEA 3323-1] 
               
               
                 (SEQ ID No: 54) 
               
               
                 MGWFMTKKYTLIYADPPWVYRDKAADGNRGAGFKYPVMSVLDICRLPVWDLADENCLLAMWWVPTQPLEALKVV 
               
               
                   
               
               
                 EAWGFRLMTMKGFTWIKCGSRQPDKLVMGMGHMTRANSEDCLFAVKGKLPTRINAGIVQSFTAPRLEHSRKPDIV 
               
               
                   
               
               
                 REKLVQLLGDVSRIELFARQTSHGFDVWGNQCEDPAVQLHPGYALDIGGLTNAFSNAPLSPTDIQGRERAA 
               
               
                   
               
               
                 &gt;AIF94871.1 Adenine DNA methyltransferase, phage-associated [ Escherichia coli   
               
               
                 O157:H7 str. SS17] 
               
               
                 (SEQ ID No: 55) 
               
               
                 MTKKYTLIYADPPWTFRDKATDGQRGASFKYPVMSLLDICRLPVWELAADNCLLAMWWVPTQPLEALKVVEAWG 
               
               
                   
               
               
                 FRLVTMKGLTWNKCGKRQTDKLVMGMGSTTRANSEDCLFAVKGNLPERINAGIIQSFTAPRLDHSRKPDMAREKL 
               
               
                   
               
               
                 VQLLGDVPRIELFARHTSHGFDVWGNQCGTPSIEMVPGIVKFLEKTNERKNDVDKGITS 
               
               
                   
               
               
                 &gt;WP_032715146.1 adenine methylase [ Klebsiella aerogenes ] 
               
               
                 (SEQ ID No: 56) 
               
               
                 MTGKYTLIYADPPWSYRDKAADGDRGAGFKYPVMNVMDICRLPVWELSADDCLLAMWWVPTQPVEALKVVEAW 
               
               
                   
               
               
                 GFRLMTMKGFTWHKINKHKGNSAIGMGHMTRANSEDCLFAVRGKLPERMDASICQHVTAPRLENSRKPDVIREKL 
               
               
                   
               
               
                 VQLLGDVPRIELFARQSSHGFDVWGNQCIAPAVELLPGCAVPVVKTEAA 
               
               
                   
               
               
                 &gt;AIA43360.1 DNA methyltransferase [ Klebsiella pneumoniae  subsp.  pneumoniae  KPNIH27] 
               
               
                 (SEQ ID No: 57) 
               
               
                 MNYDLIYCDPPWEYGNRISNGAACNHYSTMSIDDLKFLPVRKLAADNAVLAMWYTGTHNREAVELAESWGFRVRT 
               
               
                   
               
               
                 MKGFTWVKLNQNAADRFNKALSTGELVDFNDLLEMLDRETRMNGGNHTRSNTEDVLIATRGTGLPRASASVKQV 
               
               
                   
               
               
                 VHTCLGEHSAKPWEVRNRLEQLYGDVKRIELFAREEWKGWDRWGNQCNNSIEIITGLIKEVNHAA 
               
               
                   
               
               
                 &gt;WP_009320301.1 DNA methyltransferase [ Clostridioides difficile ] 
               
               
                 (SEQ ID No: 58) 
               
               
                 MPAVLFLLELHRRRKGGYKIENNQKYNIIYADPPWRYQQKRLSGAAEHHYPTMSVKDICGLKVEEIAAKDCVLFLWA 
               
               
                   
               
               
                 TFPQLPEALRVIKAWGFQYKTVAFVWLKQNKSGKGWFFGLGFWTRGNAEICLLAIKGKPHRNSNRVHQFLISPIRG 
               
               
                   
               
               
                 HSQKPEEAREKIVELMGDLPRVELFAREKTEGWDAWGNEVESDIEISSDTEKEWR 
               
               
                   
               
               
                 &gt;WP_012115592.1 MT-A70 family protein [ Xanthobacter autotrophicus ] 
               
               
                 (SEQ ID No: 59) 
               
               
                 MNGLWQFGDLKMFGYDLIVADPPWDFELYSEAGEGKSAKAHYGTMKLDEIAALRVGDLARGDCLLLLWCCEWMP 
               
               
                   
               
               
                 PAARQRVLDAWGFTYKTTIIWRKVTRAGKVRMGPGYRARTMHEPVIVATVGNPKHTPFSSVFDGVAREHSRKPEA 
               
               
                   
               
               
                 FYRMVEAAAPKAARADLFSRQRRDGWDAFGNEVEKFDQPPAEAAE 
               
               
                   
               
               
                 &gt;KFL31466.1 DNA methyltransferase [ Devosia riboflavina ] 
               
               
                 (SEQ ID No: 60) 
               
               
                 MTAWPFGAMPMFSFDVVMADPPWSFDNWSEGGNAKNAKAQYDCMPTPDIKRLPVGHLAAGDCWLWLWATYP 
               
               
                   
               
               
                 MLPDAIEVMDAWGFRYVTAGPWVKRGTSGKLAMGTGYVLRSCSEIFLIGKNGEPKTHARDVRNVLEAPRREHSRK 
               
               
                   
               
               
                 PDEAYAMAEKLFGPGRRADLFSRETRPGWTSWGNESTKFDEVAA 
               
               
                   
               
               
                 &gt;WP_016734162.1 DNA methyltransferase [ Rhizobium phaseoli ] 
               
               
                 (SEQ ID No: 61) 
               
               
                 MRLFPDLWPFGDLQPHSFDFIMADPPWKMQEWSDNGDKSKSTQSKYRLMPLDEIKAMPVLDLAAPNCLLWLWAT 
               
               
                   
               
               
                 NPMLPQALDVLHAWGFTFATAGSWMKTTRNGKQAFGTGYIFRTSNEPILIGKRGEPKTTRSVRSSFPGLAREHSR 
               
               
                   
               
               
                 KPEEGYREAERLMPRARRLELFSRTNRVGWTTWGDEVGKFGDVA 
               
               
                   
               
               
                 &gt;KFB10357.1 Adenine-specific methyltransferase [ Nitratireductor basaltis ] 
               
               
                 (SEQ ID No: 62) 
               
               
                 MHLFDWPFGDLNPHSFDLIMADPPWAFELRSDKGEGKSAQSHYKCQTLDEIKALPVLDLAAPDCLLWLWATNPML 
               
               
                   
               
               
                 PQAFEVMAAWGFTFKTAGAWGKTTVNGKLAFGTGYIFRSAHEPILIGTRGEPRTTKSVRSLIMGQVREHSRKPEEA 
               
               
                   
               
               
                 YAAAEKLIPNARRLELFSRTDRAGWEVWGDEAGKFGEAA 
               
               
                   
               
               
                 Protein sequences for phylogenetic analysis of p1 proteins 
               
               
                 &gt;XP_001009903.1 [ Tetrahymena thermophila  SB210] 
               
               
                 (SEQ ID No: 63) 
               
               
                 MSLKKGKFQHNQSKSLWNYTLSPGWREEEVKILKSALQLFGIGKWKKIMESGCLPGKSIGQIY 
               
               
                   
               
               
                 MQTQRLLGQQSLGDFMGLQIDLEAVFNQNMKKQDVLRKNNCIINTGDNPTKEERKRRIEQNR 
               
               
                   
               
               
                 KIYGLSAKQIAEIKLPKVKKHAPQYMTLEDIENEKFTNLEILTHLYNLKAEIVRRLAEQGETIAQPS 
               
               
                   
               
               
                 IIKSLNNLNHNLEQNQNSNSSTETKVTLEQSGKKKYKVLAIEETELQNGPIATNSQKKSINGKRK 
               
               
                   
               
               
                 NNRKINSDSEGNEEDISLEDIDSQESEINSEEIVEDDEEDEQIEEPSKIKKRKKNPEQESEEDDI 
               
               
                   
               
               
                 EEDQEEDELVVNEEEIFEDDDDDEDNQDSSEDDDDDED 
               
               
                   
               
               
                 &gt;EJY79729.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 64) 
               
               
                 MSSSISAAIIAGNQNKKIAESKSLWNYALSPGWTQQEVEILKIALMKFGVGRWKTIEQSQCLPT 
               
               
                   
               
               
                 KTMSQMYLQTQRLVGQQSLAEFMGLHLDLEQIFIKNAERQGAGVFRKNGCIINTGDNMTKVQI 
               
               
                   
               
               
                 AKLRKKNSKIFGLTQPFVQSLHLPKAKVKEWLKVLTLDQILSAKSNFSTAEKIHYLKILENALER 
               
               
                   
               
               
                 KLKKILRLQELVSIYRPCNIGIVVQKRLGSSIGDEYFEYVDCVKIEEKSVGNLDFALPNRNTDSTS 
               
               
                   
               
               
                 LNEDFSFLDSTQKPQKLKAGSGRENKRKKMRDGLKDERAQRQSLMEALDEQEFDETKFQDS 
               
               
                   
               
               
                 &gt;EJY78001.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 65) 
               
               
                 MSVHHKMADSKSLHNYTLSPGWTREEVDILKIALMKFGIGKWKKIQKSGCLPSKTISQMNLQT 
               
               
                   
               
               
                 QRLLGQQSLAEFMGLHVYLDRVFRDNSLKTGPEIQRKNNFIINTGNNLTQPEKEKRLRLNKQK 
               
               
                   
               
               
                 YGLDLAFIKTLRLPKPESATGGKREAILSMDQIFAQKSHFTVVEKLKHLEALKNALCSKLGKIER 
               
               
                   
               
               
                 RRRNKELSKIYRPLGQLIVVQKNADDQYEFVDIIDENE 
               
               
                   
               
               
                 &gt;ORX69504.1 [ Linderina pennispora ] 
               
               
                 (SEQ ID No: 66) 
               
               
                 MSSATPYAPRSMPTGQRNVVRSNDSASLWNCTLSPGWTQEEVQVLRKALMKFGVGNWMKII 
               
               
                   
               
               
                 ESECLPGKTIAQMNLQTQRMLGQQSTAEFNGLHLDAFVIGELNSKKQGPGIKRKNNCIVNTGG 
               
               
                   
               
               
                 KLTRDEVVKRQQKHREQYEVKAEVWRAIVLPKPDNPLILLEKKREELKKVRLELEEIMKQIEET 
               
               
                   
               
               
                 &gt;ORX78557.1 [ Basidiobolus meristosporus  CBS 931.73] 
               
               
                 (SEQ ID No: 67) 
               
               
                 MTDVYKPRSMPVGARNVLRSNDSASLWNCTLSPGWTEPEVHILRKAVMKFGIGNWAKIIESQ 
               
               
                   
               
               
                 CLFGKTIAQMNLQLQRMLGQQSTAEFAGLHLDPFVIGEINSKKQGPGIKRKNNCIVNTGGKLTR 
               
               
                   
               
               
                 EEIKRRLLEHKRTYEISEEEWRSIELPKPEDPGAVLIAKKDELKMLEDELLRVVQKIQKAREERR 
               
               
                   
               
               
                 SKSVDSSSVDGSVDDEARETKRRRK 
               
               
                   
               
               
                 &gt;EJY73777.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 68) 
               
               
                 MSHATSHGNSTEKDKKNSGNMVAESKSLWNYALSPQWTPQEVDVLKIALMKFGIGKWTIIDK 
               
               
                   
               
               
                 SGILPTKTIQQCYLQTQRILGQQSLAEFMGLHVDIDKIALDNRRKNGIRKMGFLVNQGGKLTPE 
               
               
                   
               
               
                 EKAHYQEINRQKYGLSPEEVETIKLPPPCSVEIYDINKIINPKSKLTTIEKINHCIKLQDALLEKLEN 
               
               
                   
               
               
                 IKNKKIPTGAGFSSSRVYENMRGYDPQLLLNSHVTGQLDHSMQDLTIDERYSDLDEEEDPLAM 
               
               
                   
               
               
                 ASIIDSQATPQPQKIKSSVPNKASTTPSAKEMNQIKDIIDSVIAENSAQQSKNLAQEKPKLKFSLV 
               
               
                   
               
               
                 KATESNLLQSAAQNSDDVVMEEDSKLQHIETFSTVTQTATDQSNSQSKSQNNIASDSLKDSLE 
               
               
                   
               
               
                 QNDLSKSLTDSLEMQQYSAEKKLNQAPMSKNSDKPKKKRLNKRKLPSDDEFETL 
               
               
                   
               
               
                 &gt;XP_021883515.1 [ Lobosporangium transversale ] 
               
               
                 (SEQ ID No: 69) 
               
               
                 MSSGSTPRSMTAGARNILRSNDSASLWNYTVAPGWSMKEAEILRKALMKFGIGNWSKIIESN 
               
               
                   
               
               
                 CLVGKTNAQMNLQTQRMLGQQSTAEFAGLHIDPRVIGQKNSLIQGDHIRRKNGCIVNTGAKLS 
               
               
                   
               
               
                 REEIRRRVAENKEQYELPEEEWSSIELPLPDDPHLLLEAKKSEKVRLELELKNVQRQIAMLRKV 
               
               
                   
               
               
                 GRKFETGSESPKTELDDDERDEFIEDQPLGKRARIEA 
               
               
                   
               
               
                 &gt;EJY81929.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 70) 
               
               
                 MSSSISAAIMAGNQNKKIAESKSLWNYALSPGWTQQEVEILKIALMKFGVGRWSAINKSGVLP 
               
               
                   
               
               
                 TKQIQQCYLQTQRLIGQQSLAEFMGLHLDIDRIAADNKQKRGIRKQGFLVNQGCKLTPEEKDEL 
               
               
                   
               
               
                 RKINQEKYGLSAEHVEAIKLPAPCHLVEIFQIDKIMHPRSTLSTMDKIKHLIKLEDALKSKLEMIRE 
               
               
                   
               
               
                 GKRQQKFEQLQQKLKTTEASGRGSVTRVQRQMSDLHLGSSHQNRNSDLDEENDESVMIIDE 
               
               
                   
               
               
                 SQQENLTPKGKAQAMLTHQKYNEVTQTMIKQGDDSRQQQHLPLDSTSASVSNPSSTSKSST 
               
               
                   
               
               
                 MKSNSMKQSETAIASMKPSSIGKKTKVDSSFVTKQSNQQSTAPIQKQAHQQNLDRNRSELGS 
               
               
                   
               
               
                 TFAQQASVDTQNSNNQGTSTASGNFISQSDDEEALMPKLKRRRVEDSE 
               
               
                   
               
               
                 &gt;EJY76686.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 71) 
               
               
                 MRVYLKFCNRKQIHYTHTMSSSISAAIMAGNQNKKIAESKSLWNYALSPGWTQQEVEILKIALM 
               
               
                   
               
               
                 KFGVGRWSAINKSGVLPTKQIQQCYLQTQRLIGQQSLAEFMGLHLDIDRIAADNKQKRGIRKQ 
               
               
                   
               
               
                 GFLVNQGCKLTPEEKDELRKINQEKYGLTAEHVEAIKLPAPCHLVEIFQIDKIMHPRSTLSTMDK 
               
               
                   
               
               
                 IKHLIKLEDALKSKLEMIREGKRQQKFEQLQQKLKTTEASGRGSVTRVQRQMSDLHLGSAHQN 
               
               
                   
               
               
                 RNSDLDEENDQSVMIIDESQQQNLTPKGKAQTMLTNQTQTMKKQADDSRDEQHLPLISTSAS 
               
               
                   
               
               
                 VSNPSSTSKSSALKLNSMKQSDTAIASMKPSSSGKKTKVDSSFVSKQSNQQSTSYSETNVDT 
               
               
                   
               
               
                 QNSNNQGTSTASGNFISQSDDEEALMPKLKRRRVEDSE 
               
               
                   
               
               
                 &gt;EJY80746.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 72) 
               
               
                 MRVYLKFCNRKQIHYTHTMSSSISAAIMAGNQNKKIAESKSLWNYALSPGWTQQEVEILKIALM 
               
               
                   
               
               
                 KFGVGRWSAINKSGVLPTKQIQQCYLQTQRLIGQQSLAEFMGLHLDIDRIAADNKQKRGIRKQ 
               
               
                   
               
               
                 GFLVNQGCKLTPEEKDELRKINQEKYGLTAEHVEAIKLPAPCHLVEIFQIDKIMHPRSTLSTMDK 
               
               
                   
               
               
                 IKHLIKLEDALKSKLEMIREGKRQQKFEQLQQKLKTTEASGRGSVTRVQRQMSDLHLGSAHQN 
               
               
                   
               
               
                 RNSDLDEENDQSVMIIDESQQQNLTPKGKAQTMLTNQTQTMKKQADDSREEQHLPLNSTSAS 
               
               
                   
               
               
                 VSNPSSTSKSSALKLNSMKQSDTAIASMKPSSSGKKTKVDSSFVSKQSNQQSTGPIQKQAHQ 
               
               
                   
               
               
                 QNLDRNRSELGSTFAQQTNVDTQNSNNQGTSTASGNFISQSDDEEALMPKLKRRRVKDSE 
               
               
                   
               
               
                 &gt;ORX56566.1 [ Piromyces finnis ] 
               
               
                 (SEQ ID No: 73) 
               
               
                 MSIPKPRSMPVGFRNILRPNDSTSLWNCTLSPGWTQEESDILRDALIFYGIGNWKDIIEHGCLP 
               
               
                   
               
               
                 DKTNAQMNLQLQRMLGQQSTAEFQNLHIDPYEIGKINSQKQGPNIRRKNGFIINTGGKLSREDI 
               
               
                   
               
               
                 KRKIQENKENYELPEEVWSKIVLPNREVVTINEKRQKLNKLEEELDSVLKQIVNRRRELRGMTP 
               
               
                   
               
               
                 LKETEMKSIVNRSNQNDTKTEEKEIKEEESTTVNEEKIENTETSSISIISTNENEQSENISSSSPIV 
               
               
                   
               
               
                 KSEQKKKRVVSRRKNKRRVNSDDEDFLPPGKSRSKRTRRTPKKSSN 
               
               
                   
               
               
                 &gt;ORX79686.1 [ Anaeromyces robustus ] 
               
               
                 (SEQ ID No: 74) 
               
               
                 MSIPKPRSMPTGFRNILRPNDSTSLWNCTLSPGWTQEESDILRDALIYYGIGNWKDIIEHGCLP 
               
               
                   
               
               
                 DKTNAQMNLQLQRMLGQQSTAEFQNLHIDPYVIGKINSQKQGPNIRRKNGFIINTGGKLSREDI 
               
               
                   
               
               
                 RRKIQENKENYELPKEEWSKIVLPNREVVIKNKVQEAINEKREKLNKLEDELDSVLKAIVNRRR 
               
               
                   
               
               
                 ELRGMIPLKDSEMKSLVNRSAKNEGENKTETTNNEESNNTNNSDDIKDENNETSTSSHIFTNN 
               
               
                   
               
               
                 DNELSENNSSSSSSNSISNKKKRFLRREVRRGKRRYNYDDDDFMPSGNRSRKSRKI 
               
               
                   
               
               
                 &gt;ORZ01404.1 [ Syncephalastrum racemosum ] 
               
               
                 (SEQ ID No: 75) 
               
               
                 MSNNKENNVNKPRSMTAGARNVLRSNDSTSLWNCTLSPGWTQDESEVLRKALMKFGVGNW 
               
               
                   
               
               
                 AKIIESGCLPGKTNAQMNLQLQRLLGQQSTAEFAGLHIDPKVIGEKNSKIQGPHIKRKNNCIVNT 
               
               
                   
               
               
                 GDKLSRDKLRARVMSNKEEYELPEEVWKNIELPKVKDPLMLLEGKKEEMRKLKTELEKVQAKI 
               
               
                   
               
               
                 QQLRQAQPARVQELQSQIEVARSPSPSAPDSPALSV 
               
               
                   
               
               
                 &gt;XP_001698763.1 [ Chlamydomonas reinhardtii ] 
               
               
                 (SEQ ID No: 76) 
               
               
                 MAFAAALAEKRGPRVGDAASLWNFTPAPGWSREEVQILRLCLMKHGVGQWMQILSTGLLPG 
               
               
                   
               
               
                 KLIQQLNGQTQRLLGQQSLAAYTGLKVDVDRIRVDNETRTDATRKAGLIINDGPNLTKEMKEK 
               
               
                   
               
               
                 MRQDAVAKYGLTPEQVAEVDEQLAEIAAAFNPASTSAAAGAGSGAAAAGQAAAAGSGAGGS 
               
               
                   
               
               
                 GNLMAQPTEQLSAEQLGQLLLRLRNRLACLVDRARGRAGLPPRTAPRWATEAAAAACLAAM 
               
               
                   
               
               
                 AAAEASAPQAPAAAAGGQEGAAGPVMVSVPFSREVLAEATACRVRSGTAAGARGNAPGAQ 
               
               
                   
               
               
                 GGVRKRTSKGGKAKGGDREWSPEGEENTAPQPRGGGKRKSGAVAGGEEADGVASGRAKR 
               
               
                   
               
               
                 ASRPKRGSSKHDPYVDDNDYGDEGIDPFDVGDDLDDMNPHGRYGNGGGRRADPSEAISALT 
               
               
                   
               
               
                 AMGFTQSKARGALRECNFNVELAVEWLFANCL 
               
               
                   
               
               
                 &gt;PNW76495.1 [ Chlamydomonas reinhardtii ] 
               
               
                 (SEQ ID No: 77) 
               
               
                 MAFAAALAEKRGPRVGDAASLWNFTPAPGWSREEVQILRLCLMKHGVGQWMQILSTGLLPG 
               
               
                   
               
               
                 KLIQQLNGQTQRLLGQQSLAAYTGLKVDVDRIRVDNETRTDATRKAGLIINDGPNLTKEMKEK 
               
               
                   
               
               
                 MRQDAVAKYGLTPEQVAEVDEQLAEIAAAFNPASTSAAAGAGSGAAAAGQAAAAGSGAGGS 
               
               
                   
               
               
                 GQAATAADAGGAAGRGTGSAGGAAAAAPPRNALAISTGVLAATLLDASLGNLMAQPTEQLSA 
               
               
                   
               
               
                 EQLGQLLLRLRNRLACLVDRARGRAGLPPRTAPRWATEAAAAACLAAMAAAEASAPQAPAAA 
               
               
                   
               
               
                 AGGQEGAAGPVMVSVPFSREVLAEATACRVRSGTAAGARGNAPGAQGGVRKRTSKGGKAK 
               
               
                   
               
               
                 GGDREWSPEGEENTAPQPRGGGKRKSGAVAGGEEADGVASGRAKRASRPKRGSSKHDPY 
               
               
                   
               
               
                 VDDNDYGDEGIDPFDVGDDLDDMNPHGRYGNGGGRRADPSEAISALTAMGFTQSKARGALR 
               
               
                   
               
               
                 ECNFNVELAVEWLFANCL 
               
               
                   
               
               
                 &gt;ORZ17038.1 [ Absidia repens ] 
               
               
                 (SEQ ID No: 78) 
               
               
                 MSSPSSPSPIKPRSMLTGSRNVVRSNDSASLWNCTLSPGWNEEQSETLRHAVMKYGIGNWA 
               
               
                   
               
               
                 KIIDSGYLPGKTNAQMNLQLQRLLGQQSTAEFAGLHIDPKVIGEQNSRIQGPEIRRKNNTIVNTG 
               
               
                   
               
               
                 DKLSREALRERILRNKEKYELPESVWQAIELEHVTDEDALLEEKKKTLREMKSQLKVVQRQIKN 
               
               
                   
               
               
                 LEFMHPLHAAKLKFELEKLAPSSSTSSSSSSPSPSSSSSPSSSSSKPSVSGTEEEMREAVDEE 
               
               
                   
               
               
                 RGSDEEIDELVEETDEEETSVSPKVGTRTKKVRTN 
               
               
                   
               
               
                 &gt;ORX56339.1 [ Hesseltinella vesiculosa ] 
               
               
                 (SEQ ID No: 79) 
               
               
                 MIANSTATPKPRSMKAGARNVLRSNDSASLWNCTLSPGWTEQESEILRQLAIKFGIGNWAKIIE 
               
               
                   
               
               
                 SDCLPGKTNAQMNLQLQRLLGQQSTAEFAGLHIDPKVIGEKNSKIQGPHIKRKNTTIVNTGGKL 
               
               
                   
               
               
                 SREELRERQAKNKEMYEMPKSAWDSIDLDELRDMNSLKLKKKEDKDALKKQKLTQLKTKLTK 
               
               
                   
               
               
                 SQNNLKKVQAELKQIAMVDPERVAELKKELSRASSPLSNEVSVIEESPAKKQRTS 
               
               
                   
               
               
                 &gt;ORX54764.1 [Piromyces finnis] 
               
               
                 (SEQ ID No: 80) 
               
               
                 MVVEKDLAQENKIKEELNKKHEWVKEMRKKFCVRKEFENTKNLILEDGTLNQEYFRLSKGTVL 
               
               
                   
               
               
                 KTNEVRKWTSIERNLLIKGIEKYGIGHFREISESLLPKWSGNDLRIKTIHLIGRQNLKLYKDWKG 
               
               
                   
               
               
                 GEEDIKREYNRNKEIGLKCNAWKNNCLIDDGNGKVKEMIEATEPKH 
               
               
                   
               
               
                 &gt;ORX84766.1 [ Anaeromyces robustus ] 
               
               
                 (SEQ ID No: 81) 
               
               
                 MVVEKETNKENIKNIKEELDKKHAWVKEMRKKFCVRKEFENTKILILEDGTLNQDYFRLSKGTV 
               
               
                   
               
               
                 LKTNEVRKWTSIERGLLIKGIEKYGIGHFREISENLLPKWSGNDLRIKTIHLIGRQNLKLYKDWK 
               
               
                   
               
               
                 GNEEDIKREYNRNKEIGLKCNAWKNNCLVDDGHGKVKAMIEATENN 
               
               
                   
               
               
                 &gt;ORY98423.1 [ Syncephalastrum racemosum ] 
               
               
                 (SEQ ID No: 82) 
               
               
                 MMTATDEDVDMKDVDIKLESNQETEQKILTPEEQKEKEKQDWIRQLRLKFCIRPEYEITKNMIF 
               
               
                   
               
               
                 PDGTLNQDYFRPPKGAKVEEARKWTEVEKELLIQGIEKYGIGNFGEVSKALLPAWSTNDLRIK 
               
               
                   
               
               
                 CIRLIGRQNLQLYRGWKGNADDIAREYNRNKELGLKYGTWKQGVLVYDDDGLVEKEILAQDA 
               
               
                   
               
               
                 AAKGEDVDMN 
               
               
                   
               
               
                 &gt;XP_021886199.1 [ Lobosporangium transversale ] 
               
               
                 (SEQ ID No: 83) 
               
               
                 MEINQEQLPSSSSILHPTSTSSSSSPSPSPSPASPKPERVFDARQRRINEIRLKFCIRDEFPITK 
               
               
                   
               
               
                 NMIHPDGTLNQDYFRPPRGSKPVEVARKWTDKERELLIKGIEKYGIGHFREISEEFLPLWSGN 
               
               
                   
               
               
                 DLRIKTMRLVGRQNLQLYKDWKGNEQDLAREFELNKAIGLKYGAWKAGTLVADDDGLVAKAI 
               
               
                   
               
               
                 EEQWPGSNSGTGKTTAVIGISSEENSEVSTPLNDEDVDME 
               
               
                   
               
               
                 &gt;ORY01319.1 [ Basidiobolus meristosporus  CBS 931.73] 
               
               
                 (SEQ ID No: 84) 
               
               
                 MEVDQNDSSVAKETAEQPETPEISKELLERQEWIKNMRLQFCVRPEFEVTKNIIHEDGMLNQE 
               
               
                   
               
               
                 YFLPPKGAKLEAEPERKWTETERNLLIQGIQQYGIGHFREISEALLPQWSGNDLRVKSMRLMG 
               
               
                   
               
               
                 RQNLQLYKDWKGSIEDIEREYERNKAIGLKYNTWKNSTLVYDDAGLVLKAIEASEPKP 
               
               
                   
               
               
                 &gt;ORZ26026.1 [ Absidia repens ] 
               
               
                 (SEQ ID No: 85) 
               
               
                 MAIDSLQDTEDDRTNDQNDESRESSPTPLSPEEQAQKERHEDWINQIRLKFCIRPEFEVTKNIIH 
               
               
                   
               
               
                 PDGRLNQEYFHPPKGYKPEDARKWTETEKQLLIKGIEEHGIGNFGLISKESLPKWSTNDLRVK 
               
               
                   
               
               
                 CIRLIGRQNLQLYRGWKGNADDITREYERNKEIGLKYGTWKQGVLVYDDDGMVEKELLATAAT 
               
               
                   
               
               
                 PADSMSMEEDEDMATD 
               
               
                   
               
               
                 &gt;ORX67568.1 [ Linderina pennispora ] 
               
               
                 (SEQ ID No: 86) 
               
               
                 MDTASPDDGAIAQPMLGVEDADFWRQKQEWVKQMRLQFSRRPEFPETHNMIDDEGMLNQE 
               
               
                   
               
               
                 YFQPPKDAVAPKERKWGDDEKRRLLEGIEKHGIGHFREISEESLPEWSGNDLRMKAIRLMGR 
               
               
                   
               
               
                 QNLQLYKGWKGDAAAIGLKHGTWKGGALVYDDDGVVLKAIQESNRANPP 
               
               
                   
               
               
                 &gt;XP_001699352.1 [ Chlamydomonas reinhardtii ] 
               
               
                 (SEQ ID No: 87) 
               
               
                 MAACSAACDSHVVPQPSPGSWGMPEDRDNYIVQMRRRYSPAGMLNADGSINQDFFKPRRV 
               
               
                   
               
               
                 VLVADRAKWGDAEREGLYKGLEVHGVGKWREINRDYLKGQWDDQQVRIRAARLLGSQSLVR 
               
               
                   
               
               
                 YMGWKGSKAKVDAEYAKNKAIGEATGCWKAGQLVEDDHGSVRKYFEAQQAGGEQ 
               
               
                   
               
               
                 Protein sequences for phylogenetic analysis of p2 proteins 
               
               
                 &gt;XP_001017830.3 [ Tetrahymena thermophila  SB210] 
               
               
                 (SEQ ID No: 88) 
               
               
                 MNQMGVIAIKRKQSYQLNVKINYINTAHQIKKPCQYIQKCILFRLLYKFCKQLIPLNFNLFLIFYFY 
               
               
                   
               
               
                 HLLFHLIFNYLLKFAKKINKLIRNQRKNREKKEAFKHKKIQININHYNYLKQNIQQVGIIFQNKKSK 
               
               
                   
               
               
                 LTLKLVQKKSLSEYYRKIKMKKNGKSQNQPLDFTQYAKNMRKDLSNQDICLEDGALNHSYFLT 
               
               
                   
               
               
                 KKGQYWTPLNQKALQRGIELFGVGNWKEINYDEFSGKANIVELELRTCMILGINDITEYYGKKIS 
               
               
                   
               
               
                 EEEQEEIKKSNIAKGKKENKLKDNIYQKLQQMQ 
               
               
                   
               
               
                 &gt;XP_001699352.1 [ Chlamydomonas reinhardtii ] 
               
               
                 (SEQ ID No: 89) 
               
               
                 MAACSAACDSHVVPQPSPGSWGMPEDRDNYIVQMRRRYSPAGMLNADGSINQDFFKPRRV 
               
               
                   
               
               
                 VLVADRAKWGDAEREGLYKGLEVHGVGKWREINRDYLKGQWDDQQVRIRAARLLGSQSLVR 
               
               
                   
               
               
                 YMGWKGSKAKVDAEYAKNKAIGEATGCWKAGQLVEDDHGSVRKYFEAQQAGGEQ 
               
               
                   
               
               
                 &gt;EJY77156.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 90) 
               
               
                 MSTAKQQQAQQHLLPKHSNMRVGSVSNELDYAKRNYIIKMRQSFIEVNKNIYFEDGSLNFKYF 
               
               
                   
               
               
                 NVKKGHYWSKEINEELIKGVIKYGATNYKDIKNKMEIFKKEWSETEIRLRICRLLKCYNLKVYEG 
               
               
                   
               
               
                 HKFNSREEILEQATLNKEEAIKQKKICGGILYNPPHEQDDGIMSSYFNLKNKNNTPVKASAQ 
               
               
                   
               
               
                 &gt;ORZ26026.1 [ Absidia repens ] 
               
               
                 (SEQ ID No: 91) 
               
               
                 MAIDSLQDTEDDRTNDQNDESRESSPTPLSPEEQAQKERHDWINQIRLKFCIRPEFEVTKNIIH 
               
               
                   
               
               
                 PDGRLNQEYFHPPKGYKPEDARKWTETEKQLLIKGIEEHGIGNFGLISKESLPKWSTNDLRVK 
               
               
                   
               
               
                 CIRLIGRQNLQLYRGWKGNADDITREYERNKEIGLKYGTWKQGVLVYDDDGMVEKELLATAAT 
               
               
                   
               
               
                 PADSMSMEEDEDMATD 
               
               
                   
               
               
                 &gt;ORY96423.1 [ Syncephalastrum racemosum ] 
               
               
                 (SEQ ID No: 92) 
               
               
                 MMTATDEDVDMKDVDIKLESNQETEQKILTPEEQKEKEKQDWIRQLRLKFCIRPEYEITKNMIF 
               
               
                   
               
               
                 PDGTLNQDYFRPPKGAKVEEARKWTEVEKELLIQGIEKYGIGNFGEVSKALLPAWSTNDLRIK 
               
               
                   
               
               
                 CIRLIGRQNLQLYRGWKGNADDIAREYNRNKELGLKYGTWKQGVLVYDDDGLVEKEILAQDA 
               
               
                   
               
               
                 AAKGEDVDMN 
               
               
                   
               
               
                 &gt;XP_021886199.1 [ Lobosporangium transversale ] 
               
               
                 (SEQ ID No: 93) 
               
               
                 MEINQEQLPSSSSILHPTSTSSSSSPSPSPSPASPKPERVFDARQRRINEIRLKFCIRDEFPITK 
               
               
                   
               
               
                 NMIHPDGTLNQDYFRPPRGSKPVEVARKWTDKERELLIKGIEKYGIGHFREISEEFLPLWSGN 
               
               
                   
               
               
                 DLRIKTMRLVGRQNLQLYKDWKGNEQDLAREFELNKAIGLKYGAWKAGTLVADDDGLVAKAI 
               
               
                   
               
               
                 EEQWPGSNSGTGKTTAVIGISSEENSEVSTPLNDEDVDME 
               
               
                   
               
               
                 &gt;ORY01319.1 [ Basidiobolus meristosporus  CBS 931.73] 
               
               
                 (SEQ ID No: 94) 
               
               
                 MEVDQNDSSVAKETAEQPETPEISKELLERQEWIKNMRLQFCVRPEFEVTKNIIHEDGMLNQE 
               
               
                   
               
               
                 YFLPPKGAKLEAEPERKWTETERNLLIQGIQQYGIGHFREISEALLPQWSGNDLRVKSMRLMG 
               
               
                   
               
               
                 RQNLQLYKDWKGSIEDIEREYERNKAIGLKYNTWKNSTLVYDDAQLVLKAIEASEPKP 
               
               
                   
               
               
                 &gt;ORX67568.1 [ Linderina pennispora ] 
               
               
                 (SEQ ID No: 95) 
               
               
                 MDTASPDDGAIAQPMLGVEDADFWRQKQEWVKQMRLQFSRRPEFPETHNMIDDEGMLNQE 
               
               
                   
               
               
                 YFQPPKDAVAPKERKWGDDEKRRLLEGIEKHGIGHFREISEESLPEWSGNDLRMKAIRLMGR 
               
               
                   
               
               
                 QNLQLYKGWKGDAAAIGLKHGTWKGGALVYDDDGVVLKAIQESNRANPP 
               
               
                   
               
               
                 &gt;ORX84766.1 [ Anaeromyces robustus ] 
               
               
                 (SEQ ID No: 96) 
               
               
                 MVVEKETNKENIKNIKEELDKKHAWVKEMRKKFCVRKEFENTKILILEDGTLNQDYFRLSKGTV 
               
               
                   
               
               
                 LKTNEVRKWTSIERGLLIKGIEKYGIGHFREISENLLPKWSGNDLRIKTIHLIGRQNLKLYKDWK 
               
               
                   
               
               
                 GNEEDIKREYNRNKEIGLKCNAWKNNCLVDDGHGKVKAMIEATENN 
               
               
                   
               
               
                 &gt;ORX54764.1 [ Piromyces finnis ] 
               
               
                 (SEQ ID No: 97) 
               
               
                 MVVEKDLAQENKIKEELNKKHEWVKEMRKKFCVRKEFENTKNLILEDGTLNQEYFRLSKGTVL 
               
               
                   
               
               
                 KTNEVRKWTSIERNLLIKGIEKYGIGHFREISESLLPKWSGNDLRIKTIHLIGRQNLKLYKDWKG 
               
               
                   
               
               
                 GEEDIKREYNRNKEIGLKCNAWKNNCLIDDGNGKVKEMIEATEPKH 
               
               
                   
               
               
                 &gt;ORX56334.1 [ Hesseltinella vesiculosa ] 
               
               
                 (SEQ ID No: 98) 
               
               
                 MLAGDAELVEKPHNALNAEDTEMEDVDHSSHPDTTVDLSPEQLRLQEKQAWINQMRLKFCV 
               
               
                   
               
               
                 REEFEITKNMIHPDGILNQDYFKPPKKSKKKKSKSKSKGTDETKDDTEAKGEDNKEDEDME 
               
               
                   
               
               
                 &gt;PNW76495.1 [ Chlamydomonas reinhardtii ] 
               
               
                 (SEQ ID No: 99) 
               
               
                 MAFAAALAEKRGPRVGDAASLWNFTPAPGWSREEVQILRLCLMKHGVGQWMQILSTGLLPG 
               
               
                   
               
               
                 KLIQQLNGQTQRLLGQQSLAAYTGLKVDVDRIRVDNETRTDATRKAGLIINDGPNLTKEMKEK 
               
               
                   
               
               
                 MRQDAVAKYGLTPEQVAEVDEQLAEIAAAFNPASTSAAAGAGSGAAAAGQAAAAGSGAGGS 
               
               
                   
               
               
                 GQAATAADAGGAAGRGTGSAGGAAAAAPPRNALAISTGVLAATLLDASLGNLMAQPTEQLSA 
               
               
                   
               
               
                 EQLGQLLLRLRNRLACLVDRARGRAGLPPRTAPRWATEAAAAACLAAMAAAEASAPQAPAAA 
               
               
                   
               
               
                 AGGQEGAAGPVMVSVPFSREVLAEATACRVRSGTAAGARGNAPGAQGGVRKRTSKGGKAK 
               
               
                   
               
               
                 GGDREWSPEGEENTAPQPRGGGKRKSGAVAGGEEADGVASGRAKRASRPKRGSSKHDPY 
               
               
                   
               
               
                 VDDNDYGDEGIDPFDVGDDLDDMNPHGRYGNGGGRRADPSEAISALTAMGFTQSKARGALR 
               
               
                   
               
               
                 ECNFNVELAVEWLFANCL 
               
               
                   
               
               
                 &gt;XP_001698763.1 [ Chlamydomonas reinhardtii ] 
               
               
                 (SEQ ID No: 100) 
               
               
                 MAFAAALAEKRGPRVGDAASLWNFTPAPGWSREEVQILRLCLMKHGVGQWMQILSTGLLPG 
               
               
                   
               
               
                 KLIQQLNGQTQRLLGQQSLAAYTGLKVDVDRIRVDNETRTDATRKAGLIINDGPNLTKEMKEK 
               
               
                   
               
               
                 MRQDAVAKYGLTPEQVAEVDEQLAEIAAAFNPASTSAAAGAGSGAAAAGQAAAAGSGAGGS 
               
               
                   
               
               
                 GNLMAQPTEQLSAEQLGQLLLRLRNRLACLVDRARGRAGLPPRTAPRWATEAAAAACLAAM 
               
               
                   
               
               
                 AAAEASAPQAPAAAAGGQEGAAGPVMVSVPFSREVLAEATACRVRSGTAAGARGNAPGAQ 
               
               
                   
               
               
                 GGVRKRTSKGGKAKGGDREWSPEGEENTAPQPRGGGKRKSGAVAGGEEADGVASGRAKR 
               
               
                   
               
               
                 ASRPKRGSSKHDPYVDDNDYGDEGIDPFDVGDDLDDMNPHGRYGNGGGRRADPSEAISALT 
               
               
                   
               
               
                 AMGFTQSKARGALRECNFNVELAVEWLFANCL 
               
               
                   
               
               
                 &gt;XP_011237366.1 [ Mus musculus ] 
               
               
                 (SEQ ID No: 101) 
               
               
                 MPRRQAEAMDIDAEREKITQEIQELERILYPGSTSVHFEVSESSLSSDSEADSLPDEDLETAGA 
               
               
                   
               
               
                 PILEEEGSSESSNDEEDPKDKALPEDPETCLQLNMVYQEVIREKLAEVSQLLAQNQEQQEEILF 
               
               
                   
               
               
                 DLSGTKCPKVKDGRSLPSYMYIGHFLKPYFKDKVTGVGPPANEETREKATQGIKAFEQLLVTK 
               
               
                   
               
               
                 WKHWEKALLRKSVVSDRLQRLLQPKLLKLEYLHEKQSRVSSELERQALEKQIKEAEKEIQDIN 
               
               
                   
               
               
                 QLPEEALLGNRLDSHDWEKISNINFEGARSAEEIRKFWQSSEHPSISKQEWSTEEVERLKAIA 
               
               
                   
               
               
                 ATHGHLEWHLVAEELGTSRSAFQCLQKFQQYNKTLKRKEWTEEEDHMLTQLVQEMRVGNHI 
               
               
                   
               
               
                 PYRKIVYFMEGRDSMQLIYRWTKSLDPSLKRGFWAPEEDAKLLQAVAKYGAQDWFKIREEVP 
               
               
                   
               
               
                 GRSDAQCRDRYIRRLHFSLKKGRWNAKEEQQLIQLIEKYGVGHWARIASELPHRSGSQCLSK 
               
               
                   
               
               
                 WKILARKKQHLQRKRGQRPRHSSQWSSSGSSSSSSEDYGSSSGSDGSSGSENSDVELEAS 
               
               
                   
               
               
                 LEKSRALTPQQYRVPDIDLWVPTRLITSQSQREGTGCYPQHPAVSCCTQDASQNHHKEGSTT 
               
               
                   
               
               
                 VSAAEKNQLQVPYETHSTVPRGDRFLHFSDTHSASLKDPACKPVLKVPLEKMPKLIRTRPPTQ 
               
               
                   
               
               
                 SHTLMKERPKQPLLPSSRSGSDPGNNTAGPHLRQLWHGTYQNKQRRKRQALHRRLLKHRLL 
               
               
                   
               
               
                 LAVIPWVGDINLACTQAPRRPATVQTKADSIRMQLECARLASTPVFTLLIQLLQIDTAGMEVV 
               
               
                   
               
               
                 RERKSQPPALLQPGTRNTQPHLLQASSNAKNNTGCLPSMTGEQTAKRASHKGRPRLGSCRT 
               
               
                   
               
               
                 EATPFQVPVAAPRGLRPKPKTVSELLREKRLRESHAKKATQALGLNSQLLVSSPVILQPPLLPV 
               
               
                   
               
               
                 PHGSPVVGPATSSVELSVPVAPVMVSSSPSGSWPVGGISATDKQPPNLQTISLNPPHKGTQV 
               
               
                   
               
               
                 AAPAAFRSLALAPGQVPTGGHLSTLGQTSTTSQKQSLPKVLPILRAAPSLTQLSVQPPVSGQP 
               
               
                   
               
               
                 LATKSSLPVNWVLTTQKLLSVQVPAVVGLPQSVMTPETIGLQAKQLPSPAKTPAFLEQPPAST 
               
               
                   
               
               
                 DTEPKGPQGQEIPPTPGPEKAALDLSLLSQESEAAIVTWLKGCQGAFVPPLGSRMPYHPPSL 
               
               
                   
               
               
                 CSLRALSSLLLQKQDLEQKASSLAASQAAGAQPDPKAGALQASLELVQRQFRDNPAYLLLKTR 
               
               
                   
               
               
                 FLAIFSLPAFLATLPPNSIPTTLSPDVAVVSESDSEDLGDLELKDRARQLDCMACRVQASPAAP 
               
               
                   
               
               
                 DPVQSHLVSPGQRAPSPGEVSAPSPLDASDGLDDLNVLRTRRARHSRR 
               
               
                   
               
               
                 &gt;XP_006497966.1 [ Mus musculus ] 
               
               
                 (SEQ ID No: 102) 
               
               
                 MPRRQAEAMDIDAEREKITQEIQELERILYPGSTSVHFEVSESSLSSDSEADSLPDEDLETAGA 
               
               
                   
               
               
                 PILEEEGSSESSNDEEDPKDKALPEDPETCLQLNMVYQEVIREKLAEVSQLLAQNQEQQEEILF 
               
               
                   
               
               
                 DLSGTKCPKVKDGRSLPSYMYIGHFLKPYFKDKVTGVGPPANEETREKATQGIKAFEQLLVTK 
               
               
                   
               
               
                 WKHWEKALLRKSVVSDRLQRLLQPKLLKLEYLHEKQSRVSSELERQALEKQIKEAEKEIQDIN 
               
               
                   
               
               
                 QLPEEALLGNRLDSHDWEKISNINFEGARSAEEIRKFWQSSEHPSISKQEWSTEEVERLKAIA 
               
               
                   
               
               
                 ATHGHLEWHLVAEELGTSRSAFQCLQKFQQYNKTLKRKEWTEEEDHMLTQLVQEMRVGNHI 
               
               
                   
               
               
                 PYRKIVYFMEGRDSMQLIYRWTKSLDPSLKRGFWAPEEDAKLLQAVAKYGAQDWFKIREEVP 
               
               
                   
               
               
                 GRSDAQCRDRYIRRLHFSLKKGRWNAKEEQQLIQLIEKYGVGHWARIASELPHRSGSQCLSK 
               
               
                   
               
               
                 WKILARKKQHLQRKRGQRPRHSSQWSSSGSSSSSSEDYGSSSGSDGSSGSENSDVELEAS 
               
               
                   
               
               
                 LEKSRALTPQQYRVPDIDLWVPTRLITSQSQREGTGCYPQHPAVSCCTQDASQNHHKEGSTT 
               
               
                   
               
               
                 VSAAEKNQLQVPYETHSTVPRGDRFLHFSDTHSASLKDPACKSHTLMKERPKQPLLPSSRSG 
               
               
                   
               
               
                 SDPGNNTAGPHLRQLWHGTYQNKQRRKRQALHRRLLKHRLLLAVIPWVGDINLACTQAPRRP 
               
               
                   
               
               
                 ATVQTKADSIRMQLECARLASTPVFTLLIQLLQIDTAGCMEVVRERKSQPPALLQPGTRNTQP 
               
               
                   
               
               
                 HLLQASSNAKNNTGCLPSMTGEQTAKRASHKGRPRLGSCRTEATPFQVPVAAPRGLRPKPK 
               
               
                   
               
               
                 TVSELLREKRLRESHAKKATQALGLNSQLLVSSPVILQPPLLPVPHGSPVVGPATSSVELSVPV 
               
               
                   
               
               
                 APVMVSSSPSGSWPVGGISATDKQPPNLQTISLNPPHKGTQVAAPAAFRSLALAPGQVPTGG 
               
               
                   
               
               
                 HLSTLGQTSTTSQKQSLPKVLPILRAAPSLTQLSVQPPVSGQPLATKSSLPVNWVLTTQKLLSV 
               
               
                   
               
               
                 QVPAVVGLPQSVMTPETIGLQAKQLPSPAKTPAFLEQPPASTDTEPKGPQGQEIPPTPGPEKA 
               
               
                   
               
               
                 ALDLSLLSQESEAAIVTWLKGCQGAFVPPLGSRMPYHPPSLCSLRALSSLLLQKQDLEQKASS 
               
               
                   
               
               
                 LAASQAAGAQPDPKAGALQASLELVQRQFRDNPAYLLLKTRFLAIFSLPAFLATLPPNSIPTTLS 
               
               
                   
               
               
                 PDVAVVSESDSEDLGDLELKDRARQLDCMACRVQASPAAPDPVQSHLVSPGQRAPSPGEVS 
               
               
                   
               
               
                 APSPLDASDGLDDLNVLRTRRARHSRR 
               
               
                   
               
               
                 &gt;EJY86254.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 103) 
               
               
                 MSVHHKMADSKSLHNYTLSPGWTREEVDILKIALMKFGIGKWKKIQKSGCLPSKTISQMNLQT 
               
               
                   
               
               
                 QRLLGQQSLAEFMGLHVYLDRVFRDNSLKTGPEIQRKNNFIINTGNNLTQPEKEKRLRLNKQK 
               
               
                   
               
               
                 YGLDLAFIKTLRLPKPESATGGKREAILSMDQIFAQKSHFTVVEKLKHLEALKNALCSKLGKIER 
               
               
                   
               
               
                 RRRNKELSKIYRPLCQLIVVQKNADDQYEFVDIIDENE 
               
               
                   
               
               
                 &gt;ORX69504.1 [ Linderina pennispora ] 
               
               
                 (SEQ ID No: 104) 
               
               
                 MSSATPYAPRSMPTGQRNVVRSNDSASLWNCTLSPGWTQEEVQVLRKALMKFGVGNWMKII 
               
               
                   
               
               
                 ESECLPGKTIAQMNLQTQRMLGQQSTAEFNGLHLDAFVIGELNSKKQGPGIKRKNNCIVNTGG 
               
               
                   
               
               
                 KLTRDEVVKRQQKHREQYEVKAEVWRAIVLPKPDNPLILLEKKREELKKVRLELEEIMKQIEET 
               
               
                   
               
               
                 EKLVDVPEHAPGTKRARE 
               
               
                   
               
               
                 &gt;NP_003077.2 [ Homo sapiens ] 
               
               
                 (SEQ ID No: 105) 
               
               
                 MDVDAEREKITQEIKELERILDPGSSGSHVEISESSLESDSEADSLPSEDLDPADPPISEEERW 
               
               
                   
               
               
                 GEASNDEDDPKDKTLPEDPETCLQLNMVYQEVIQEKLAEANLLLAQNREQQEELMRDLAGSK 
               
               
                   
               
               
                 GTKVKDGKSLPPSTYMGHFMKPYFKDKVTGVGPPANEDTREKAAQGIKAFEELLVTKWKNW 
               
               
                   
               
               
                 EKALLRKSVVSDRLQRLLQPKLLKLEYLHQKQSKVSSELERQALEKQGREAEKEIQDINQLPEE 
               
               
                   
               
               
                 ALLGNRLDSHDWEKISNINFEGSRSAEEIRKFWQNSEHPSINKQEWSREEEERLQAIAAAHGH 
               
               
                   
               
               
                 LEWQKIAEELGTSRSAFQCLQKFQQHNKALKRKEWTEEEDRMLTQLVQEMRVGSHIPYRRIV 
               
               
                   
               
               
                 YYMEGRDSMQLIYRWTKSLDPGLKKGYWAPEEDAKLLQAVAKYGEQDWFKIREEVPGRSDA 
               
               
                   
               
               
                 QCRDRYLRRLHFSLKKGRWNLKEEEQLIELIEKYGVGHWAKIASELPHRSGSQCLSKWKIMM 
               
               
                   
               
               
                 GKKQGLRRRRRRARHSVRWSSTSSSGSSSGSSGGSSSSSSSSSEEDEPEQAQAGEGDRAL 
               
               
                   
               
               
                 LSPQYMVPDMDLWVPARQSTSQPWRGGAGAWLGGPAASLSPPKGSSASQGGSKEASTTA 
               
               
                   
               
               
                 AAPGEETSPVQVPARAHGPVPRSAQASHSADTRPAGAEKQALEGGRRLLTVPVETVLRVLRA 
               
               
                   
               
               
                 NTAARSCTQKEQLRQPPLPTSSPGVSSGDSVARSHVQWLRHRATQSGQRRWRHALHRRLL 
               
               
                   
               
               
                 NRRLLLAVTPWVGDVVVPCTQASQRPAVVQTQADGLREQLQQARLASTPVFTLFTQLFHIDT 
               
               
                   
               
               
                 AGCLEVVRERKALPPRLPQAGARDPPVHLLQASSSAQSTPGHLFPNVPAQEASKSASHKGSR 
               
               
                   
               
               
                 RLASSRVERTLPQASLLASTGPRPKPKTVSELLQEKRLQEARAREATRGPVVLPSQLLVSSSVI 
               
               
                   
               
               
                 LQPPLPHTPHGRPAPGPTVLNVPLSGPGAPAAAKPGTSGSWQEAGTSAKDKRLSTMQALPL 
               
               
                   
               
               
                 APVFSEAEGTAPAASQAPALGPGQISVSCPESGLGQSQAPAASRKQGLPEAPPFLPAAPSPT 
               
               
                   
               
               
                 PLPVQPLSLTHIGGPHVATSVPLPVTWVLTAQGLLPVPVPAVVSLPRPAGTPGPAGLLATLLPP 
               
               
                   
               
               
                 LTETRAAQGPRAPALSSSWQPPANMNREPEPSCRTDTPAPPTHALSQSPAEADGSVAFVPG 
               
               
                   
               
               
                 EAQVAREIPEPRTSSHADPPEAEPPWSGRLPAFGGVIPATEPRGTPGSPSGTQEPRGPLGLE 
               
               
                   
               
               
                 KLPLRQPGPEKGALDLEKPPLPQPGPEKGALDLGLLSQEGEAATQQWLGGQRGVRVPLLGS 
               
               
                   
               
               
                 RLPYQPPALCSLRALSGLLLHKKALEHKATSLVVGGEAERPAGALQASLGLVRGQLQDNPAYL 
               
               
                   
               
               
                 LLRARFLAAFTLPALLATLAPQGVRTTLSVPSRVGSESEDEDLLSELELADRDGQPGCTTATC 
               
               
                   
               
               
                 PIQGAPDSGKCSASSCLDTSNDPDDLDVLRTRHARHTRKRRRLV 
               
               
                   
               
               
                 &gt;XP_016870547.1 [ Homo sapiens ] 
               
               
                 (SEQ ID No: 106) 
               
               
                 MDVDAEREKITQEIKELERILDPGSSGSHVEISESSLESDSEADSLPSEDLDPADPPISEEERW 
               
               
                   
               
               
                 GEASNDEDDPKDKTLPEDPETCLQLNMVYQEVIQEKLAEANLLLAQNREQQEELMRDLAGSK 
               
               
                   
               
               
                 GTKVKDGKSLPPSTYMGHFMKPYFKDKVTGVGPPANEDTREKAAQGIKAFEELLVTKWKNW 
               
               
                   
               
               
                 EKALLRKSVVSDRLQRLLQPKLLKLEYLHQKQSKVSSELERQALEKQGREAEKEIQDINQLPEE 
               
               
                   
               
               
                 ALLGNRLDSHDWEKISNINFEGSRSAEEIRKFWQNSEHPSINKQEWSREEEERLQAIAAAHGH 
               
               
                   
               
               
                 LEWQKIAEELGTSRSAFQCLQKFQQHNKALKRKEWTEEEDRMLTQLVQEMRVGSHIPYRRIV 
               
               
                   
               
               
                 YYMEGRDSMQLIYRWTKSLDPGLKKGYWAPEEDAKLLQAVAKYGEQDWFKIREEVPGRSDA 
               
               
                   
               
               
                 QCRDRYLRRLHFSLKKGRWNLKEEEQLIELIEKYGVGHWAKIASELPHRSGSQCLSKWKIMM 
               
               
                   
               
               
                 GKKQGLRRRRRRARHSVRWSSTSSSGSSSGSSGGSSSSSSSSSEEDEPEQAQAGEGDRAL 
               
               
                   
               
               
                 LSPQYMVPDMDLWVPARQSTSQPWRGGAGAWLGGPAASLSPPKGSSASQGGSKEASTTA 
               
               
                   
               
               
                 AAPGEETSPVQVPARAHGPVPRSAQASHSADTRPAGAEKQALEGGRRLLTVPVETVLRVLRA 
               
               
                   
               
               
                 NTAARSCTQWLRHRATQSGQRRWRHALHRRLLNRRLLLAVTPWVGDVVVPCTQASQRPAV 
               
               
                   
               
               
                 VQTQADGLREQLQQARLASTPVFTLFTQLFHIDTAGCLEVVRERKALPPRLPQAGARDPPVHL 
               
               
                   
               
               
                 LQASSSAQSTPGHLFPNVPAQEASKSASHKGSRRLASSRVERTLPQASLLASTGPRPKPKTV 
               
               
                   
               
               
                 SELLQEKRLQEARAREATRGPVVLPSQLLVSSSVILQPPLPHTPHGRPAPGPTVLNVPLSGPG 
               
               
                   
               
               
                 APAAAKPGTSGSWQEAGTSAKDKRLSTMQALPLAPVFSEAEGTAPAASQAPALGPGQISVSC 
               
               
                   
               
               
                 PESGLGQSQAPAASRKQGLPEAPPFLPAAPSPTPLPVQPLSLTHIGGPHVATSVPLPVTWVLT 
               
               
                   
               
               
                 AQGLLPVPVPAVVSLPRPAGTPGPAGLLATLLPPLTETRAAQGPRAPALSSSWQPPANMNRE 
               
               
                   
               
               
                 PEPSCRTDTPAPPTHALSQSPAEADGSVAFVPGEAQVAREIPEPRTSSHADPPEAEPPWSGR 
               
               
                   
               
               
                 LPAFGGVIPATEPRGTPGSPSGTQEPRGPLGLEKLPLRQPGPEKGALDLEKPPLPQPGPEKG 
               
               
                   
               
               
                 ALDLGLLSQEGEAATQQWLGGQRGVRVPLLGSRLPYQPPALCSLRALSGLLLHKKALEHKAT 
               
               
                   
               
               
                 SLVVGGEAERPAGALQASLGLVRGQLQDNPAYLLLRARFLAAFTLPALLATLAPQGVRTTLSV 
               
               
                   
               
               
                 PSRVGSESEDEDLLSELELADRDGQPGCTTATCPIQGAPDSGKCSASSCLDTSNDPDDLDVL 
               
               
                   
               
               
                 RTRHARHTRKRRRLV 
               
               
                   
               
               
                 &gt;XP_020936800.1 [ Sus scrofa ] 
               
               
                 (SEQ ID No: 107) 
               
               
                 MDVDAEREKISKEIKELERILDPGSSGINDDVSESSLDSDSEAESLPDDDADATGPLLSEDERW 
               
               
                   
               
               
                 GDASNDEDDAKERALPEDPETCLQLNMVYQEVVREKLAEVSLLLAQNREQQEEVSWALAGS 
               
               
                   
               
               
                 GGRRVKDGRSPPARLYVGHFMKPYFKDKVTGAGPPANEDTREKAAQGVKAFEELLVTKWKS 
               
               
                   
               
               
                 WEKALLRKAVVSDRLQRLLQPKLLKLEYLQQKQSRATSDAERQALEKQVREAEKEVQDISQL 
               
               
                   
               
               
                 PEEALLGHRLDSHDWEKIANVNFEGGRSAEETRKFWQNHEHPSINKQEWSAQEVDRLKAIAA 
               
               
                   
               
               
                 KHGHLRWQEIAEELGTRRSAFQCLQKYQQHNAALKRREWTQEEDRMLTQLVQAMGVGSHIP 
               
               
                   
               
               
                 YRRIAYYMEGRDSTQLIYRWTKSLDPALKKGLWAPEEDAKLLQAVAKYGEQDWFKIREEVPG 
               
               
                   
               
               
                 RSDAQCRDRYLRRLRLSLKKGRWSAQEEERLLELIGKHGVGHWAKIASELPHRTDSQCLSK 
               
               
                   
               
               
                 WKIMARKQQSRGRRRRRPLRRVCWSSSSEDSEDSGDSGGSSSSSSSSEDVEPEGAPEARA 
               
               
                   
               
               
                 DGPAPPSAQHPVPDMDLWVPTRQSARVPWGVGPGAWPGHRSASPRPPEGSDVAPGEEAG 
               
               
                   
               
               
                 RAQAPSETPSASLRGGGCPRSADARPSGSEGLADEGPRRPLTVPLETVLRVLRTNTAALCRA 
               
               
                   
               
               
                 LKEKLRRPRLLGSPLGPSPSDGSVARPRVQPRWRRRHALQRRLLERQLLMAVSPWVGDVTL 
               
               
                   
               
               
                 PCAPWRPAVLHRRADGIGKQLQGARLASTPVFTLLIQLFRIDTAGCMEVVRERRAQPPALPSG 
               
               
                   
               
               
                 GRVPSSARNSPGHLFQNGSARGAAKKSASHSGGGGPQSAPAPSGPRPKPKTVSELLREKRL 
               
               
                   
               
               
                 REARARKAAQGPAVLPPQGLLSSPAILQPLPPQQLPVSGAVLSGPGGPAVASPGAPGPWAS 
               
               
                   
               
               
                 AKEGPPSLHALALAPASMAAGVTPAAPRAPALGPSQVPASCHLSSLGQSQAPATSRKQGLPE 
               
               
                   
               
               
                 APPFLPAAPSPIQLPVQPRSLTPALAAHTGASHVVASTPLPVTWVLTAQGLLPVPAVVGLPRP 
               
               
                   
               
               
                 AGPPDPEGLSGTPPPSLTETRAGRGPKQPPAHVSVGPDPPAKTPPTAQSPAEGDGDVAHGP 
               
               
                   
               
               
                 GGPSCPGEAQVAGEASVPRTLSPAKPLADHPEAEPCGSSQLPLPGGLSPGGAPTRHQGLER 
               
               
                   
               
               
                 PPPPWPGPEKGAPDLRLLSQESEAAVRGWLTGQRGVCVPPLASRLPYQPPTLCSLRALSGLL 
               
               
                   
               
               
                 LHKKALEHRAASLVPSGAAGAQQAPLGQVRERLQSSPAYLLLKARFLAAFALPALLATLPPHG 
               
               
                   
               
               
                 VPTTLSAAAGVDSESDDDSLDELELADNGGPLGGWPSGRQAGPAAPTPTQGAPGEGSAAP 
               
               
                   
               
               
                 GLDSDDLDILRTRHAWHARKRRRLV 
               
               
                   
               
               
                 &gt;XP_021883515.1 [ Lobosporangium transversale ] 
               
               
                 (SEQ ID No: 108) 
               
               
                 MSSGSTPRSMTAGARNILRSNDSASLWNYTVAPGWSMKEAEILRKALMKFGIGNWSKIIESN 
               
               
                   
               
               
                 CLVGKTNAQMNLQTQRMLGQQSTAEFAGLHIDPRVIGQKNSLIQGDHIRRKNGCIVNTGAKLS 
               
               
                   
               
               
                 REEIRRRVAENKEQYELPEEEWSSIELPLPDDPHLLLEAKKSEKVRLELELKNVQRQIAMLRKV 
               
               
                   
               
               
                 GRKFETGSESPKTELDDDERDEFIEDQPLGKRARIEA 
               
               
                   
               
               
                 &gt;EJY76686.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 109) 
               
               
                 MRVYLKFCNRKQIHYTHTMSSSISAAIMAGNQNKKIAESKSLWNYALSPGWTQQEVEILKIALM 
               
               
                   
               
               
                 KFGVGRWSAINKSGVLPTKQIQQCYLQTQRLIGQQSLAEFMGLHLDIDRIAADNKQKRGIRKQ 
               
               
                   
               
               
                 GFLVNQGCKLTPEEKDELRKINQEKYGLTAEHVEAIKLPAPCHLVEIFQIDKIMHPRSTLSTMDK 
               
               
                   
               
               
                 IKHLIKLEDALKSKLEMIREGKRQQKFEQLQQKLKTTEASGRGSVTRVQRQMSDLHLGSAHQN 
               
               
                   
               
               
                 RNSDLDEENDQSVMIIDESQQQNLTPKGKAQTMLTNQTQTMKKQADDSRDEQHLPLISTSAS 
               
               
                   
               
               
                 VSNPSSTSKSSALKLNSMKQSDTAIASMKPSSSGKKTKVDSSFVSKQSNQQSTSYSETNVDT 
               
               
                   
               
               
                 QNSNNQGTSTASGNFISQSDDEEALMPKLKRRRVEDSE 
               
               
                   
               
               
                 &gt;EJY73777.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 110) 
               
               
                 MSHATSHGNSTEKDKKNSGNMVAESKSLWNYALSPGWTPQEVDVLKIALMKFGIGKWTIIDK 
               
               
                   
               
               
                 SGILPTKTIQQCYLQTQRILGQQSLAEFMGLHVDIDKIALDNRRKNGIRKMGFLVNQGGKLTPE 
               
               
                   
               
               
                 EKAHYQEINRQKYGLSPEEVETIKLPPPCSVEIYDINKIINPKSKLTTIEKINHCIKLQDALLEKLEN 
               
               
                   
               
               
                 IKNKKIPTGAGFSSSRVYENMRGYDPQLLLNSHVTGQLDHSMQDLTIDERYSDLDEEEDPLAM 
               
               
                   
               
               
                 ASIIDSQATPQPQKIKSSVPNKASTTPSAKEMNQIKDIIDSVIAENSAQQSKNLAQEKPKLKFSLV 
               
               
                   
               
               
                 KATESNLLQSAAQNSDDVVMEEDSKLQHIETFSTVTQTATDQSNSQSKSQNNIASDSLKDSLE 
               
               
                   
               
               
                 QNDLSKSLTDSLEMQQYSAEKKLNQAPMSKNSDKPKKKRLNKRKLPSDDEFETL 
               
               
                   
               
               
                 &gt;EJY79729.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 111) 
               
               
                 MSSSISAAIIAGNQNKKIAESKSLWNYALSPGWTQQEVEILKIALMKFGVGRWKTIEQSQCLPT 
               
               
                   
               
               
                 KTMSQMYLQTQRLVGQQSLAEFMGLHLDLEQIFIKNAERQGAGVFRKNGCIINTGDNMTKVQI 
               
               
                   
               
               
                 AKLRKKNSKIFGLTQPFVQSLHLPKAKVKEWLKVLTLDQILSAKSNFSTAEKIHYLKILENALER 
               
               
                   
               
               
                 KLKKILRLQELVSIYRPCNIGIVVQKRLGSSIGDEYFEYVDCVKIEEKSVGNLDFALPNRNTDSTS 
               
               
                   
               
               
                 LNEDFSFLDSTQKPQKLKAGSGRENKRKKMRDGLKDERAQRQSLMEALDEQEFDETKFQDS 
               
               
                   
               
               
                 DGEMPDLNM 
               
               
                   
               
               
                 &gt;EJY81929.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 112) 
               
               
                 MSSSISAAIMAGNQNKKIAESKSLWNYALSPGWTQQEVEILKIALMKFGVGRWSAINKSGVLP 
               
               
                   
               
               
                 TKQIQQCYLQTQRLIGQQSLAEFMGLHLDIDRIAADNKQKRGIRKQGFLVNQGCKLTPEEKDEL 
               
               
                   
               
               
                 RKINQEKYGLSAEHVEAIKLPAPCHLVEIFQIDKIMHPRSTLSTMDKIKHLIKLEDALKSKLEMIRE 
               
               
                   
               
               
                 GKRQQKFEQLQQKLKTTEASGRGSVTRVQRQMSDLHLGSSHQNRNSDLDEENDESVMIIDE 
               
               
                   
               
               
                 SQQENLTPKGKAQAMLTHQKYNEVTQTMIKQGDDSRQQQHLPLDSTSASVSNPSSTSKSST 
               
               
                   
               
               
                 MKSNSMKQSETAIASMKPSSIGKKTKVDSSFVTKQSNQQSTAPIQKQAHQQNLDRNRSELGS 
               
               
                   
               
               
                 TFAQQASVDTQNSNNQGTSTASGNFISQSDDEEALMPKLKRRRVEDSE 
               
               
                   
               
               
                 &gt;EJY80746.1 [ Oxytricha trifallax ] 
               
               
                 (SEQ ID No: 113) 
               
               
                 MRVYLKFCNRKQIHYTHTMSSSISAAIMAGNQNKKIAESKSLWNYALSPGWTQQEVEILKIALM 
               
               
                   
               
               
                 KFGVGRWSAINKSGVLPTKQIQQCYLQTQRLIGQQSLAEFMGLHLDIDRIAADNKQKRGIRKQ 
               
               
                   
               
               
                 GFLVNQGCKLTPEEKDELRKINQEKYGLTAEHVEAIKLPAPCHLVEIFQIDKIMHPRSTLSTMDK 
               
               
                   
               
               
                 IKHLIKLEDALKSKLEMIREGKRQQKFEQLQQKLKTTEASGRGSVTRVQRQMSDLHLGSAHQN 
               
               
                   
               
               
                 RNSDLDEENDQSVMIIDESQQQNLTPKGKAQTMLTNQTQTMKKQADDSREEQHLPLNSTSAS 
               
               
                   
               
               
                 VSNPSSTSKSSALKLNSMKQSDTAIASMKPSSSGKKTKVDSSFVSKQSNQQSTGPIQKQAHQ 
               
               
                   
               
               
                 QNLDRNRSELGSTFAQQTNVDTQNSNNQGTSTASGNFISQSDDEEALMPKLKRRRVKDSE 
               
               
                   
               
               
                 &gt;ORX78557.1 [ Basidiobolus meristosporus  CBS 931.73] 
               
               
                 (SEQ ID No: 114) 
               
               
                 MTDVYKPRSMPVGARNVLRSNDSASLWNCTLSPGWTEPEVHILRKAVMKFGIGNWAKIIESQ 
               
               
                   
               
               
                 CLFGKTIAQMNLQLQRMLGQQSTAEFAGLHLDPFVIGEINSKKQGPGIKRKNNCIVNTGGKLTR 
               
               
                   
               
               
                 EEIKRRLLEHKRTYEISEEEWRSIELPKPEDPGAVLIAKKDELKMLEDELLRVVQKIQKAREERR 
               
               
                   
               
               
                 SKSVDSSSVDGSVDDEARETKRRRK 
               
               
                   
               
               
                 &gt;ORX79686.1 [ Anaeromyces robustus ] 
               
               
                 (SEQ ID No: 115) 
               
               
                 MSIPKPRSMPTGFRNILRPNDSTSLWNCTLSPGWTQEESDILRDALIYYGIGNWKDIIEHGCLP 
               
               
                   
               
               
                 DKTNAQMNLQLQRMLGQQSTAEFQNLHIDPYVIGKINSQKQGPNIRRKNGFIINTGGKLSREDI 
               
               
                   
               
               
                 RRKIQENKENYELPKEEWSKIVLPNREVVIKNKVQEAINEKREKLNKLEDELDSVLKAIVNRRR 
               
               
                   
               
               
                 ELRGMIPLKDSEMKSLVNRSAKNEGENKTETTNNEESNNTNNSDDIKDENNETSTSSHIFTNN 
               
               
                   
               
               
                 DNELSENNSSSSSSNSISNKKKRFLRREVRRGKRRYNYDDDDFMPSGNRSRKSRKI 
               
               
                   
               
               
                 &gt;ORX56566.1 [ Piromyces finnis ] 
               
               
                 (SEQ ID No: 116) 
               
               
                 MSIPKPRSMPVGFRNILRPNDSTSLWNCTLSPGWTQEESDILRDALIFYGIGNWKDIIEHGCLP 
               
               
                   
               
               
                 DKTNAQMNLQLQRMLGQQSTAEFQNLHIDPYEIGKINSQKQGPNIRRKNGFIINTGGKLSREDI 
               
               
                   
               
               
                 KRKIQENKENYELPEEVWSKIVLPNREVVTINEKRQKLNKLEEELDSVLKQIVNRRRELRGMTP 
               
               
                   
               
               
                 LKETEMKSIVNRSNQNDTKTEEKEIKEEESTTVNEEKIENTETSSISIISTNENEQSENISSSSPIV 
               
               
                   
               
               
                 KSEQKKKRVVSRRKNKRRVNSDDEDFLPPGKSRSKRTRRTPKKSSN 
               
               
                   
               
               
                 &gt;XP_001009903.1 [ Tetrahymena thermophila  SB210] 
               
               
                 (SEQ ID No: 117) 
               
               
                 MSLKKGKFQHNQSKSLWNYTLSPGWREEEVKILKSALQLFGIGKWKKIMESGCLPGKSIGQIY 
               
               
                   
               
               
                 MQTQRLLGQQSLGDFMGLQIDLEAVFNQNMKKQDVLRKNNCIINTGDNPTKEERKRRIEQNR 
               
               
                   
               
               
                 KIYGLSAKQIAEIKLPKVKKHAPQYMTLEDIENEKFTNLEILTHLYNLKAEIVRRLAEQGETIAQPS 
               
               
                   
               
               
                 IIKSLNNLNHNLEQNQNSNSSTETKVTLEQSGKKKYKVLAIEETELQNGPIATNSQKKSINGKRK 
               
               
                   
               
               
                 NNRKINSDSEGNEEDISLEDIDSQESEINSEEIVEDDEEDEQIEEPSKIKKRKKNPEQESEEDDI 
               
               
                   
               
               
                 EEDQEEDELVVNEEEIFEDDDDDEDNQDSSEDDDDDED 
               
               
                   
               
               
                 &gt;XP_020936799.1 [ Sus scrofa ] 
               
               
                 (SEQ ID No: 118) 
               
               
                 MDVDAEREKISKEIKELERILDPGSSGINDDVSESSLDSDSEAESLPDDDADATGPLLSEDERW 
               
               
                   
               
               
                 GDASNDEDDAKERALPEDPETCLQLNMVYQEVVREKLAEVSLLLAQNREQQEEVSWALAGS 
               
               
                   
               
               
                 GGRRVKDGRSPPARLYVGHFMKPYFKDKVTGAGPPANEDTREKAAQGVKAFEELLVTKWKS 
               
               
                   
               
               
                 WEKALLRKAVVSDRLQRLLQPKLLKLEYLQQKQSRATSDAERQALEKQVREAEKEVQDISQL 
               
               
                   
               
               
                 PEEALLGHRLDSHDWEKIANVNFEGGRSAEETRKFWQNHEHPSINKQEWSAQEVDRLKAIAA 
               
               
                   
               
               
                 KHGHLRWQEIAEELGTRRSAFQCLQKYQQHNAALKRREWTQEEDRMLTQLVQAMGVGSHIP 
               
               
                   
               
               
                 YRRIAYYMEGRDSTQLIYRWTKSLDPALKKGLWAPEEDAKLLQAVAKYGEQDWFKIREEVPG 
               
               
                   
               
               
                 VTFEARAFPASRQRTSLPCAPLWPPALWVSRLGNRRGGRQPRGFSRTPRSVCRRYLRRLRL 
               
               
                   
               
               
                 SLKKGRWSAQEEERLLELIGKHGVGHWAKIASELPHRTDSQCLSKWKIMARKQQSRGRRRR 
               
               
                   
               
               
                 RPLRRVCWSSSSEDSEDSGDSGGSSSSSSSSEDVEPEGAPEARADGPAPPSAQHPVPDMD 
               
               
                   
               
               
                 LWVPTRQSARVPWGVGPGAWPGHRSASPRPPEGSDVAPGEEAGRAQAPSETPSASLRGG 
               
               
                   
               
               
                 GCPRSADARPSGSEGLADEGPRRPLTVPLETVLRVLRTNTAALCRALKEKLRRPRLLGSPLGP 
               
               
                   
               
               
                 SPSDGSVARPRVQPRWRRRHALQRRLLERQLLMAVSPWVGDVTLPCAPWRPAVLHRRADG 
               
               
                   
               
               
                 IGKQLQGARLASTPVFTLLIQLFRIDTAGCMEVVRERRAQPPALPSGGRVPSSARNSPGHLFQ 
               
               
                   
               
               
                 NGSARGAAKKSASHSGGGGPQSAPAPSGPRPKPKTVSELLREKRLREARARKAAQGPAVLP 
               
               
                   
               
               
                 PQGLLSSPAILQPLPPQQLPVSGAVLSGPGGPAVASPGAPGPWASAKEGPPSLHALALAPAS 
               
               
                   
               
               
                 MAAGVTPAAPRAPALGPSQVPASCHLSSLGQSQAPATSRKQGLPEAPPFLPAAPSPIQLPVQ 
               
               
                   
               
               
                 PRSLTPALAAHTGASHVVASTPLPVTWVLTAQGLLPVPAVVGLPRPAGPPDPEGLSGTPPPSL 
               
               
                   
               
               
                 TETRAGRGPKQPPAHVSVGPDPPAKTPPTAQSPAEGDGDVAHGPGGPSCPGEAQVAGEAS 
               
               
                   
               
               
                 VPRTLSPAKPLADHPEAEPCGSSQLPLPGGLSPGGAPTRHQGLERPPPPWPGPEKGAPDLR 
               
               
                   
               
               
                 LLSQESEAAVRGWLTGQRGVCVPPLASRLPYQPPTLCSLRALSGLLLHKKALEHRAASLVPS 
               
               
                   
               
               
                 GAAGAQQAPLGQVRERLQSSPAYLLLKARFLAAFALPALLATLPPHGVPTTLSAAAGVDSESD 
               
               
                   
               
               
                 DDSLDELELADNGGPLGGWPSGRQAGPAAPTPTQGAPGEGSAAPGLDSDDLDILRTRHAWH 
               
               
                   
               
               
                 ARKRRRLV 
               
               
                   
               
               
                 &gt;XP_009300052.1 [ Danio rerio ] 
               
               
                 (SEQ ID No: 119) 
               
               
                 MKCLSVNMTHLSRDSWLYTHDVQVTYNSFIKVSPCPKMASDDLRAQRDKIQREILALESTLGA 
               
               
                   
               
               
                 DSSIADQLSSDNSSDYESDDSGPTVKRVERDDLETERLRIQREIEELENALGADAALENVLQD 
               
               
                   
               
               
                 SDHDTDSSEDSADDLELPQNVETCLQMNLVYQEVLKEKLAELEQLLIENQQQQKEIEVQLSGP 
               
               
                   
               
               
                 GNSIFSVPGVPPQKQFLGYFLKPYFKDKLTGLGPPANEETKERMKHGSIPVDNLKIKRWEGW 
               
               
                   
               
               
                 QKTLLTNAVARDTMKRMLQPKLSKMEYLSNKLCRAEGEEKEQLKAQIELIEKQIAEIRTLKDDQ 
               
               
                   
               
               
                 LLGDLQDDHDWDKISNIDFEGLRQADDLKRFWQNFLHPSINKSVWKQDEIYKLQAVAEEFKM 
               
               
                   
               
               
                 CHWDKIAEALGTNRTAFMCFQTYQRYISKTFRRTHWTEEEDDLLRELVEKMRIGNFIPYIQMS 
               
               
                   
               
               
                 HFMVGRDGSQLAYRWTSVLDPSLKKGPWSKEEDQLLRNAVAKYGTREWGRIRTEVPGRTD 
               
               
                   
               
               
                 SACRDRYLDCLRETVKKGTWSYAEMELLKEKVAKYGVGKWAKIASEIPNRVDAQCLHKWKL 
               
               
                   
               
               
                 MTRSKKPLKRPLSSITTSYPRNKRQKLLKTVKEEMFFNSSSDDESQINYMNSDESDDLAEDEN 
               
               
                   
               
               
                 LEIPQKEYVQTEMKEWIPRNAMVWTITPGSFRTLWVRLPTNEEELRESTKESGLGSDSSENS 
               
               
                   
               
               
                 ACPNDEPIMERNTILDRFGDVERTYVGMNTVVLHRRTDDEKAMFKVCMSDVKQFIQMKATEF 
               
               
                   
               
               
                 AVKKKKKIKNKKRTLRDVFSLNTDLQKAVIPWIGNVIISTPANEAIFCEGDIVGIKAASIRLQKTSV 
               
               
                   
               
               
                 FTFFIKAFHVDVNGCRTVIEIHKKLDIKMPLAINGNPKPTPISTSPKTVAVLLQQSKAASEHKKPA 
               
               
                   
               
               
                 EPSQQPSLPPSQKPSLPPAQQPTQPPSLPPSVPPSQQPTLPPPSQPSQPPPQPPSLPPSQPP 
               
               
                   
               
               
                 AQQPPQQPSLPPPQPPSLPPPQPPSLPTSQQQSLPPSQQHSLPPFQNPSLPPSQQPSLPPS 
               
               
                   
               
               
                 KQPPQPLPVRQITTPTLIYPNNLVITNPNMEGEVQHLVFKGLLLPQQPSKAVSHIPLPVMQPKT 
               
               
                   
               
               
                 PAQPIVVSKSPSVQDSNSVKSSKRICKPTKKAQALMEQSKVKSRKKEPQKQNQGNKNVVFPT 
               
               
                   
               
               
                 VTLQTSPVIKILSPARLVQVTGLSPNFSSNQTINMPDKSLTIKSPQPCSSGNLHQSAPVVVHSS 
               
               
                   
               
               
                 TNPTFVHSSVSNVSRDNLNVSSTINISPRVSRDALNPTSFLNSTTFPLPQNLSVQQSVQIVPQIP 
               
               
                   
               
               
                 INVVHKATCTKAAKTSSDSSSDESVVKQHQLSPSTGRSIPPAVFNIQPNPSTPPTLSSGPVIFN 
               
               
                   
               
               
                 PNNKVVAPKLCGLNVSSSQLPTVSTQKTKYRPIRPLGPLPVVAPPSRKVTSMSRIRAQSEGEP 
               
               
                   
               
               
                 LISLRDLPAAGVNFDSHLIFPEKSSEVDDWMDGKGGIPLPHLDTSLPYLPPSAATIKTMTDLLRA 
               
               
                   
               
               
                 KQPLLLAAKKVLPAQYQDECNEEVEVEAIRKVVAERFASNPAYLLCKARFLSCFTLPALLATINP 
               
               
                   
               
               
                 CEERQLLSEDDEEDDHLATINPSEEHQSSTEDDEEDLQTNERSQPPTARTELNMNENEASAK 
               
               
                   
               
               
                 QFSGIGPKRQRNQRIKRLIK 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 2 
               
             
            
               
                   
               
               
                 Primer sequences. 
               
            
           
           
               
               
               
            
               
                 Name 
                 Sequence (5′ to 3′) 
                 Description 
               
               
                   
               
               
                 1781.0_qF24 
                 ACTAGTCTTAAATATGAGAAAGATGATTTGAA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 TAAGAT (SEQ ID No: 120) 
                 primers 
               
               
                   
               
               
                 1781.0_qR24 
                 ATCCTAGCAATATTATCTACTTATAATTCTATT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 GACTATTAG (SEQ ID No: 121) 
                 primers 
               
               
                   
               
               
                 1781.0_qF23 
                 CTAATTAACTAATAGTCAATAGAATTATAAGT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 AGATAATATTGCT (SEQ ID No: 122) 
                 primers 
               
               
                   
               
               
                 1781.0_qR23 
                 CATTAAATCATTAACAGAGTAATGTCGTCATA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 TATTTGTC (SEQ ID No: 123) 
                 primers 
               
               
                   
               
               
                 1781.0_qF22 
                 TTTAGTGAGCATAGACAAATATATGACGACAT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 TACTC (SEQ ID No: 124) 
                 primers 
               
               
                   
               
               
                 1781.0_qR22 
                 GCGGAGATGTCTTTTTGACCTTTTGATAG 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 125) 
                 primers 
               
               
                   
               
               
                 1781.0_qF21 
                 ATGTTAACATGCTTATTATTACTATCAAAAGG 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 TCAA (SEQ ID No: 126) 
                 primers 
               
               
                   
               
               
                 1781.0_qR21 
                 GGCTGCTACTGATATTTATGTTCTTTATGTTT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 A (SEQ ID No: 127) 
                 primers 
               
               
                   
               
               
                 1781.0_qF20 
                 CAAAGAACACGAAGCTCATAAACATAAAGAA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 CAT (SEQ ID No: 128) 
                 primers 
               
               
                   
               
               
                 1781.0_qR20 
                 TGGAGCAAATGCTGCTAATAACGAG (SEQ ID 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 No: 129) 
                 primers 
               
               
                   
               
               
                 1781.0_qF19 
                 ACCTCCAGCAGCTCCGTTTCTATTATTTG 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 130) 
                 primers 
               
               
                   
               
               
                 1781.0_qR19 
                 GGCCTGGGTATTTTCCCTGCTTTA (SEQ ID 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 No: 131) 
                 primers 
               
               
                   
               
               
                 1781.0_qF18 
                 CTTCCCAGGTAAAATTTAAGGTAAATAAAGCA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 GG (SEQ ID No: 132) 
                 primers 
               
               
                   
               
               
                 1781.0_qR18 
                 TCAAGGTGGAGGACTCTTCGGTAAC (SEQ ID 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 No: 133) 
                 primers 
               
               
                   
               
               
                 1781.0_qF17 
                 ATTACGAACCCACTACCTGAATTATTGTTACC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 G (SEQ ID No: 134) 
                 primers 
               
               
                   
               
               
                 1781.0_qR17 
                 AAACGTCCTGCAGGACAACGC (SEQ ID No: 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 135) 
                 primers 
               
               
                   
               
               
                 1781.0_qF16 
                 TTGATTGAAGTTTTAATTTGGTACTGGGC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 136) 
                 primers 
               
               
                   
               
               
                 1781.0_qR16 
                 TTGGATGCTGATCTGTTTTGTTTAGAAAG 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 137) 
                 primers 
               
               
                   
               
               
                 1781.0_qF15 
                 TTGGGATTTCTTAACTGGATTTCTTTCTAAAC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 138) 
                 primers 
               
               
                   
               
               
                 1781.0_qR15 
                 CTGCTTAAATTAAGTACTTCTATGTTTGAAAT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 TAATGTTC (SEQ ID No: 139) 
                 primers 
               
               
                   
               
               
                 1781.0_qF14 
                 CAATTAAAACACGTTGAACATTAATTTCAAAC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 ATAG (SEQ ID No: 140) 
                 primers 
               
               
                   
               
               
                 1781.0_qR14 
                 TGAGGATCCAAGGTAAATTTCATACAATC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 141) 
                 primers 
               
               
                   
               
               
                 1781.0_qF13 
                 GACTGCATGTATATGCTAATGATTGTATGAAA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 TTTAC (SEQ ID No: 142) 
                 primers 
               
               
                   
               
               
                 1781.0_qR13 
                 AGTGGCATTTCCAAGGAAACATTAATAC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 143) 
                 primers 
               
               
                   
               
               
                 1781.0_qF12 
                 CAGTGTTTCCCTTTGTGTAAATGGG (SEQ ID 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 No: 144) 
                 primers 
               
               
                   
               
               
                 1781.0_qR12 
                 TCAGTGGATAAACTAGCCTAAGGAAACAC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 145) 
                 primers 
               
               
                   
               
               
                 1781.0_qF11 
                 TTTTACAGACTGGACACAGTAGTGTTTCC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 146) 
                 primers 
               
               
                   
               
               
                 1781.0_qR11 
                 CCAGTGGTATCAACATGCGGTCATC (SEQ ID 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 No: 147) 
                 primers 
               
               
                   
               
               
                 1781.0_qF10 
                 GATATATACACTCCCAGCAGTAAAGATGACC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 148) 
                 primers 
               
               
                   
               
               
                 1781.0_qR10 
                 GAATAGGCTCACTCTAAATTCGAGTGC (SEQ 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 ID No: 149) 
                 primers 
               
               
                   
               
               
                 1781.0_qF9 
                 ATTCGCTAGGTCTAAGCAAATATTGCAC 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 150) 
                 primers 
               
               
                   
               
               
                 1781.0_qR9 
                 TAAATAGCCAAAACAACCAATAAAATTAACAA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 TAACCTC (SEQ ID No: 151) 
                 primers 
               
               
                   
               
               
                 1781.0_qF8 
                 CTTTTTGAGGGCGAGGTTATTGTTAATTTTAT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 TG (SEQ ID No: 152) 
                 primers 
               
               
                   
               
               
                 1781.0_qR8 
                 GATCCATTAATTACAGAAATAAATAATAGGCA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 GCATA (SEQ ID No: 153) 
                 primers 
               
               
                   
               
               
                 1781.0_qF7 
                 ATATTGCCTGAATTATTATGCTGCCTATTATT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 TATT (SEQ ID No: 154) 
                 primers 
               
               
                   
               
               
                 1781.0_qR7 
                 AAATGTGCACCGTCATCAAATACC (SEQ ID 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 No: 155) 
                 primers 
               
               
                   
               
               
                 1781.0_qF6 
                 GGATCACTATAATCATCTGGATGACTATTGG 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 (SEQ ID No: 156) 
                 primers 
               
               
                   
               
               
                 1781.0_qR6 
                 AAGTGTAATGTAGTTTCAATGGTAGTGATGT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 G (SEQ ID No: 157) 
                 primers 
               
               
                   
               
               
                 1781.0_qF5 
                 TGACTTCTTCCAGTGGATTCACATC (SEQ ID 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 No: 158) 
                 primers 
               
               
                   
               
               
                 1781.0_qR5 
                 GCCAATTAATTCATTTGTTCGTAGAGATATGT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 AA (SEQ ID No: 159) 
                 primers 
               
               
                   
               
               
                 1781.0_qF4 
                 CACTTTATAATAAATAAGAATTATTACATATCT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 CTACGAACAA (SEQ ID No: 160) 
                 primers 
               
               
                   
               
               
                 1781.0_qR4 
                 CTCACCAGTAATTTGCAGACACC (SEQ ID 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 No: 161) 
                 primers 
               
               
                   
               
               
                 1781.0_qF3 
                 GGCTGACTGGGGTTGAGTTAATC (SEQ ID 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 No: 162) 
                 primers 
               
               
                   
               
               
                 1781.0_qR3 
                 AATATAAACAAAATGGAATATACAAAACTTGA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 ATAAGAAATAG (SEQ ID No: 163) 
                 primers 
               
               
                   
               
               
                 1781.0_qF2 
                 GAGACTGAGGATCTATTTCTTATTCAAGTTTT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 G (SEQ ID No: 164) 
                 primers 
               
               
                   
               
               
                 1781.0_qR2 
                 ATTAATACATTATTAACTTAAATATAAATATTT 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 AAAGAATTATGAACAATAAT (SEQ ID 
                 primers 
               
               
                   
                 No: 165) 
                   
               
               
                   
               
               
                 1781.0_qF1 
                 CATTTTGTTTATATTATTGTTCATAATTCTTTA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 AATATTTATATTTAAGTTAAT(SEQ ID 
                 primers 
               
               
                   
                 No: 166) 
                   
               
               
                   
               
               
                 1781.0_qR1 
                 ACAAGATAACATTGCTAATTTTCAATAAATTA 
                 Contig1781.0 tiling qPCR 
               
               
                   
                 AATTAATACATT (SEQ ID No: 167) 
                 primers 
               
               
                   
               
               
                 1781.0_F 
                 CCCCAAAACCCCAAAACCCCACTAGTCTTAA 
                 Primer pair for amplifying 
               
               
                   
                 ATATGAGAAAGATGATTTGAATAAG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 168) 
                 mini-genome 
               
               
                 1781.0_R 
                 CCCCAAAACCCCAAAACCCCACAAGATAACA 
                   
               
               
                   
                 TTGCTAATTTTCAATAAATTAAAT (SEQ ID 
                   
               
               
                   
                 No: 169) 
                   
               
               
                   
               
               
                 15118.0_F 
                 CCCCAAAACCCCAAAACCCCGATTTATGAAA 
                 Primer pair for amplifying 
               
               
                   
                 GTGCTGTATTATTAAGGAATG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 170) 
                 mini-genome 
               
               
                 15118.0_R 
                 CCCCAAAACCCCAAAACCCCATTATTCCTAC 
                   
               
               
                   
                 TTTTAGCTATATTAGAAATTCG (SEQ ID No: 
                   
               
               
                   
                 171) 
                   
               
               
                   
               
               
                 1339.1_F 
                 CCCCAAAACCCCAAAACCCCATGATGATACA 
                 Primer pair for amplifying 
               
               
                   
                 TAGATTCATTAAAATAAAAAAAAG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 172) 
                 mini-genome 
               
               
                 1339.1_R 
                 CCCCAAAACCCCAAAACCCCTTAGATGAATT 
                   
               
               
                   
                 AAATAAAGAATTCAAATAAATAC (SEQ ID 
                   
               
               
                   
                 No: 173) 
                   
               
               
                   
               
               
                 20718.0_F 
                 CCCCAAAACCCCAAAACCCCATGAATCTGAA 
                 Primer pair for amplifying 
               
               
                   
                 ATCGGGCAGTTGAATACG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 174) 
                 mini-genome 
               
               
                 20718.0_R 
                 CCCCAAAACCCCAAAACCCCATTTATCATAAT 
                   
               
               
                   
                 TATAGAGAAGATAGTGATGC (SEQ ID No: 
                   
               
               
                   
                 175) 
                   
               
               
                   
               
               
                 20822.0_F 
                 CCCCAAAACCCCAAAACCCCATGAGAGTTTG 
                 Primer pair for amplifying 
               
               
                   
                 TGAAAAATTAAGTTTG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 176) 
                 mini-genome 
               
               
                 20822.0_R 
                 CCCCAAAACCCCAAAACCCCTATATTAAATAT 
                   
               
               
                   
                 CAAGAAAAAGTAAAAAGACAG (SEQ ID No: 
                   
               
               
                   
                 177) 
                   
               
               
                   
               
               
                 21162.0_F 
                 CCCCAAAACCCCAAAACCCCAAGTCTCATTT 
                 Primer pair for amplifying 
               
               
                   
                 TGGTTAGTGATGTTTGGATTG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 178) 
                 mini-genome 
               
               
                 21162.0_R 
                 CCCCAAAACCCCAAAACCCCGTATGATCGAT 
                   
               
               
                   
                 GAATACAAAATCAAGTTGGAAG (SEQ ID No: 
                   
               
               
                   
                 179) 
                   
               
               
                   
               
               
                 11991.0_F 
                 CCCCAAAACCCCAAAACCCCACTTAAAAGGA 
                 Primer pair for amplifying 
               
               
                   
                 TTGCATGATTGTAAGGGAAATGTG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 180) 
                 mini-genome 
               
               
                 11991.0_R 
                 CCCCAAAACCCCAAAACCCCAATAATCGCAC 
                   
               
               
                   
                 TTACATTATATCTGGAGAAATG (SEQ ID No: 
                   
               
               
                   
                 181) 
                   
               
               
                   
               
               
                 5079.0_F 
                 CCCCAAAACCCCAAAACCCCTTCTACTAAATT 
                 Primer pair for amplifying 
               
               
                   
                 TCATTGATTTTTTTCAATTTC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 182) 
                 mini-genome 
               
               
                 5079.0_R 
                 CCCCAAAACCCCAAAACCCCATTTGATAGAA 
                   
               
               
                   
                 TAGAAGAGAAATTATGGAATG (SEQ ID No: 
                   
               
               
                   
                 183) 
                   
               
               
                   
               
               
                 13665.0_F 
                 CCCCAAAACCCCAAAACCCCAAGTATAAATA 
                 Primer pair for amplifying 
               
               
                   
                 AGGGAGTTGATATATAATATACTT (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 184) 
                 mini-genome 
               
               
                 13665.0_R 
                 CCCCAAAACCCCAAAACCCCATGAGAATTCC 
                   
               
               
                   
                 TATTCAAAAATGAAAAAGTAGATTG (SEQ ID 
                   
               
               
                   
                 No: 185) 
                   
               
               
                   
               
               
                 22365.0_F 
                 CCCCAAAACCCCAAAACCCCATAAGGTAGTA 
                 Primer pair for amplifying 
               
               
                   
                 TATTTTTATTAAGGATTGGAAATTA (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 186) 
                 mini-genome 
               
               
                 22365.0_R 
                 CCCCAAAACCCCAAAACCCCATAAGACTAAA 
                   
               
               
                   
                 TTTATTGAAATTATCTTGTTAATAG (SEQ ID 
                   
               
               
                   
                 No: 187) 
                   
               
               
                   
               
               
                 21620.0_F 
                 CCCCAAAACCCCAAAACCCCTTGAGCCAATA 
                 Primer pair for amplifying 
               
               
                   
                 CTGAAAAGGATGATAGTGAATAGTG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 188) 
                 mini-genome 
               
               
                 21620.0_R 
                 CCCCAAAACCCCAAAACCCCTCATTTTTTAAA 
                   
               
               
                   
                 TTGGATAGTAAGAAAAATTATAATAAAG (SEQ 
                   
               
               
                   
                 ID No: 189) 
                   
               
               
                   
               
               
                 15049.0_F 
                 CCCCAAAACCCCAAAACCCCAAGGAATAAAA 
                 Primer pair for amplifying 
               
               
                   
                 TTCAATTCCAAAATGTAAGGTGAG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 190) 
                 mini-genome 
               
               
                 15049.0_R 
                 CCCCAAAACCCCAAAACCCCGTTAAAAGAAC 
                   
               
               
                   
                 CAAGTGATATATTATAAGCCA (SEQ ID No: 
                   
               
               
                   
                 191) 
                   
               
               
                   
               
               
                 16562.0_F 
                 CCCCAAAACCCCAAAACCCCTTTATCAATTAT 
                 Primer pair for amplifying 
               
               
                   
                 AAATAAAAAGTTTTAAGTCTATTTTTAA (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 192) 
                 mini-genome 
               
               
                 16562.0_R 
                 CCCCAAAACCCCAAAACCCCATAAGACAAAT 
                   
               
               
                   
                 GCAACTTTATAAAGTAAATAAATTATC (SEQ 
                   
               
               
                   
                 ID No: 193) 
                   
               
               
                   
               
               
                 22360.0_F 
                 CCCCAAAACCCCAAAACCCCAATGCAACATT 
                 Primer pair for amplifying 
               
               
                   
                 TACTTTTAACATTAGAGATTATC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 194) 
                 mini-genome 
               
               
                 22360.0_R 
                 CCCCAAAACCCCAAAACCCCATAAGAGCAAA 
                   
               
               
                   
                 AGTTAATATAAAAATTCAAGGTG (SEQ ID 
                   
               
               
                   
                 No: 195) 
                   
               
               
                   
               
               
                 15836.0_F 
                 CCCCAAAACCCCAAAACCCCGATTTGCACAG 
                 Primer pair for amplifying 
               
               
                   
                 TTAATTTGAATTTGGTATTTG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 196) 
                 mini-genome 
               
               
                 15836.0_R 
                 CCCCAAAACCCCAAAACCCCTCATTTTTAGTA 
                   
               
               
                   
                 TTTTAAATATCATTTAGTTTTAAGTAA (SEQ 
                   
               
               
                   
                 ID No: 197) 
                   
               
               
                   
               
               
                 2324.0_F 
                 CCCCAAAACCCCAAAACCCCTTGATTGATTC 
                 Primer pair for amplifying 
               
               
                   
                 CTGAATACAAATGAAATAATATAAAG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 198) 
                 mini-genome 
               
               
                 2324.0_R 
                 CCCCAAAACCCCAAAACCCCAAGACCAAAAT 
                   
               
               
                   
                 AAAGAGGAATAATGAGAAGTAC (SEQ ID No: 
                   
               
               
                   
                 199) 
                   
               
               
                   
               
               
                 22404.0_F 
                 CCCCAAAACCCCAAAACCCCATGTAGAATTA 
                 Primer pair for amplifying 
               
               
                   
                 ATATGAGAACATCATTTTTTAAGC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 200) 
                 mini-genome 
               
               
                 22404.0_R 
                 CCCCAAAACCCCAAAACCCCATAATGTAAGA 
                   
               
               
                   
                 AATCTGATACAATAGAGAGATAAAC (SEQ ID 
                   
               
               
                   
                 No: 201) 
                   
               
               
                   
               
               
                 15403.0_F 
                 CCCCAAAACCCCAAAACCCCGAATGGAAAAT 
                 Primer pair for amplifying 
               
               
                   
                 TTGTATGAAGTTCAGAGAGAAAG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 202) 
                 mini-genome 
               
               
                 15403.0_R 
                 CCCCAAAACCCCAAAACCCCATAAGATTATC 
                   
               
               
                   
                 AGTTATAAAAATTGATAGGGGATG (SEQ ID 
                   
               
               
                   
                 No: 203) 
                   
               
               
                   
               
               
                 17795.0_F 
                 CCCCAAAACCCCAAAACCCCATCATACGATA 
                 Primer pair for amplifying 
               
               
                   
                 TCTTAAGTGTTGATCTGAATTAAAT (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 204) 
                 mini-genome 
               
               
                 17795.0_R 
                 CCCCAAAACCCCAAAACCCCGTTAGGTTTAA 
                   
               
               
                   
                 GAGTAGAAATAAAAGGAGATAAG (SEQ ID 
                   
               
               
                   
                 No: 205) 
                   
               
               
                   
               
               
                 11141.0_F 
                 CCCCAAAACCCCAAAACCCCTCTCACTATCT 
                 Primer pair for amplifying 
               
               
                   
                 TTTGTAAAAAGTTGGTAGAT (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 206) 
                 mini-genome 
               
               
                 11141.0_R 
                 CCCCAAAACCCCAAAACCCCGTTGGTTTAGA 
                   
               
               
                   
                 ATAAAGAATTGTATTAACCAAATTTAT (SEQ 
                   
               
               
                   
                 ID No: 207) 
                   
               
               
                   
               
               
                 22342.0_F 
                 CCCCAAAACCCCAAAACCCCGTGAATTAAAA 
                 Primer pair for amplifying 
               
               
                   
                 TATAAACGAATAAGATATAAAGATTG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 208) 
                 mini-genome 
               
               
                 22342.0_R 
                 CCCCAAAACCCCAAAACCCCTTAATTACTGA 
                   
               
               
                   
                 ATTGTTTATTATAAGATTATAAG (SEQ ID 
                   
               
               
                   
                 No: 209) 
                   
               
               
                   
               
               
                 2240.0_F 
                 CCCCAAAACCCCAAAACCCCGTAATGAATAA 
                 Primer pair for amplifying 
               
               
                   
                 ATTGTAAAGGTAAATTGCAA (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 210) 
                 mini-genome 
               
               
                 2240.0_R 
                 CCCCAAAACCCCAAAACCCCAATGGCAAACA 
                   
               
               
                   
                 TTTAAAATAAATATTAATATAAATTAC (SEQ 
                   
               
               
                   
                 ID No: 211) 
                   
               
               
                   
               
               
                 3531.0_F 
                 CCCCAAAACCCCAAAACCCCTAAAAGGAAAA 
                 Primer pair for amplifying 
               
               
                   
                 CAAATAGAAGAAACTGAA (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 212) 
                 mini-genome 
               
               
                 3531.0_R 
                 CCCCAAAACCCCAAAACCCCATTTGGATATT 
                   
               
               
                   
                 ATGATTAGCAGTTTAGTG (SEQ ID No: 
                   
               
               
                   
                 213) 
                   
               
               
                   
               
               
                 4701.0_F 
                 CCCCAAAACCCCAAAACCCCTTTAAATAAAAA 
                 Primer pair for amplifying 
               
               
                   
                 TCGCATGAATTAAATGCAAG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 214) 
                 mini-genome 
               
               
                 4701.0_R 
                 CCCCAAAACCCCAAAACCCCTAGGTAAATGC 
                   
               
               
                   
                 AAATTGGAGAATTTCCAATAG (SEQ ID No: 
                   
               
               
                   
                 215) 
                   
               
               
                   
               
               
                 20883.0_F 
                 CCCCAAAACCCCAAAACCCCATATTAAGAAT 
                 Primer pair for amplifying 
               
               
                   
                 TGTGTAATTTTTGAGTAAATTG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 216) 
                 mini-genome 
               
               
                 20883.0_R 
                 CCCCAAAACCCCAAAACCCCATTTAGTAGAA 
                   
               
               
                   
                 TCTTCAATAAATAAGCGTTATTG (SEQ ID 
                   
               
               
                   
                 No: 217) 
                   
               
               
                   
               
               
                 15191.0_F 
                 CCCCAAAACCCCAAAACCCCTAGCATTAAAT 
                 Primer pair for amplifying 
               
               
                   
                 TTGTAAAAAGAATGAAATTTAATAT (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 218) 
                 mini-genome 
               
               
                 15191.0_R 
                 CCCCAAAACCCCAAAACCCCAATATACATGA 
                   
               
               
                   
                 TTTTAGATAAACAACAAATAAT (SEQ ID No: 
                   
               
               
                   
                 219) 
                   
               
               
                   
               
               
                 19342.0_F 
                 CCCCAAAACCCCAAAACCCCATCAAGAATGG 
                 Primer pair for amplifying 
               
               
                   
                 ATTAGAATTTTTAATGCTTTGC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 220) 
                 mini-genome 
               
               
                 19342.0_R 
                 CCCCAAAACCCCAAAACCCCGAGGAACTAG 
                   
               
               
                   
                 GGATTACTCATTTTACTTCAG (SEQ ID No: 
                   
               
               
                   
                 221) 
                   
               
               
                   
               
               
                 15245.0_F 
                 CCCCAAAACCCCAAAACCCCATGCATGTAAT 
                 Primer pair for amplifying 
               
               
                   
                 TTTCTGTCAAAATTGAGTAAATAG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 222) 
                 mini-genome 
               
               
                 15245.0_R 
                 CCCCAAAACCCCAAAACCCCGTAAGCTAAAT 
                   
               
               
                   
                 AAGTAGACTAAATAGGTAG (SEQ ID 
                   
               
               
                   
                 No: 223) 
                   
               
               
                   
               
               
                 6109.0_F 
                 CCCCAAAACCCCAAAACCCCAACCGCAAATA 
                 Primer pair for amplifying 
               
               
                   
                 GAATATATAAAGGATAATTTA (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 224) 
                 mini-genome 
               
               
                 6109.0_R 
                 CCCCAAAACCCCAAAACCCCGAAGTACTAAA 
                   
               
               
                   
                 AATAAAAAGTAAAGTATTAAAATAAAATC 
                   
               
               
                   
                 (SEQ ID No: 225) 
                   
               
               
                   
               
               
                 22610.0_F 
                 CCCCAAAACCCCAAAACCCCGTAGACAGATT 
                 Primer pair for amplifying 
               
               
                   
                 TTCCAGTTTATAGCTGTGTTTG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 226) 
                 mini-genome 
               
               
                 22610.0_R 
                 CCCCAAAACCCCAAAACCCCTTTATGAATTTT 
                   
               
               
                   
                 CTTAAATCTGTAAATAAATAAAATAAT (SEQ 
                   
               
               
                   
                 ID No: 227) 
                   
               
               
                   
               
               
                 11875.0_F 
                 CCCCAAAACCCCAAAACCCCGTATGTTAATT 
                 Primer pair for amplifying 
               
               
                   
                 TTATGCTTTAAATGATAGTTTA (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 228) 
                 mini-genome 
               
               
                 11875.0_R 
                 CCCCAAAACCCCAAAACCCCTGGATTCCATT 
                   
               
               
                   
                 TTGAAGAATAATTTATTAAC (SEQ ID 
                   
               
               
                   
                 No: 229) 
                   
               
               
                   
               
               
                 15329.0_F 
                 CCCCAAAACCCCAAAACCCCTTGTTTCGATT 
                 Primer pair for amplifying 
               
               
                   
                 ATATTCAAAATAGGAAATTTAG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 230) 
                 mini-genome 
               
               
                 15329.0_R 
                 CCCCAAAACCCCAAAACCCCATGAATTTCAA 
                   
               
               
                   
                 TAACTTTTTATGAAAATGAATTTA (SEQ ID 
                   
               
               
                   
                 No: 231) 
                   
               
               
                   
               
               
                 20179.0_F 
                 CCCCAAAACCCCAAAACCCCTAGGAAGAAAA 
                 Primer pair for amplifying 
               
               
                   
                 TCTTGTGTGCAATTTGAGATTAAC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 232) 
                 mini-genome 
               
               
                 20179.0_R 
                 CCCCAAAACCCCAAAACCCCTTGATAAAAAC 
                   
               
               
                   
                 ATAGATTAAATACTAGTGTATAAA (SEQ ID 
                   
               
               
                   
                 No: 233) 
                   
               
               
                   
               
               
                 9936.0_F 
                 CCCCAAAACCCCAAAACCCCATATGGAATAT 
                 Primer pair for amplifying 
               
               
                   
                 TTAATTTGATTTAAATGAAACGAAATA (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 234) 
                 mini-genome 
               
               
                 9936.0_R 
                 CCCCAAAACCCCAAAACCCCTTGTAACAGTA 
                   
               
               
                   
                 AATAGAATATTTTAATTACCAAAAC (SEQ ID 
                   
               
               
                   
                 No: 235) 
                   
               
               
                   
               
               
                 16267.0_F 
                 CCCCAAAACCCCAAAACCCCTCATTTTAGAA 
                 Primer pair for amplifying 
               
               
                   
                 TTATCTGTACTTAATTATTTTG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 236) 
                 mini-genome 
               
               
                 16267.0_R 
                 CCCCAAAACCCCAAAACCCCATGAGCATGTT 
                   
               
               
                   
                 ATTTTACTTCATTAGTCAATTTG (SEQ ID 
                   
               
               
                   
                 No: 237) 
                   
               
               
                   
               
               
                 4488.0_F 
                 CCCCAAAACCCCAAAACCCCATGAAATGAAT 
                 Primer pair for amplifying 
               
               
                   
                 TCTAAGATTGAATTGCATG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 238) 
                 mini-genome 
               
               
                 4488.0_R 
                 CCCCAAAACCCCAAAACCCCAGAAGAGATCA 
                   
               
               
                   
                 ATAAATTGAGAAGGAATTG (SEQ ID 
                   
               
               
                   
                 No: 239) 
                   
               
               
                   
               
               
                 8551.0_F 
                 CCCCAAAACCCCAAAACCCCGTGTTACAATT 
                 Primer pair for amplifying 
               
               
                   
                 TGCGTTTGAAATAGTTGGTTGATA (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 240) 
                 mini-genome 
               
               
                 8551.0_R 
                 CCCCAAAACCCCAAAACCCCATATGGTAAAA 
                   
               
               
                   
                 ATTGAAGAAAGAAATTCAAGAGAA (SEQ ID 
                   
               
               
                   
                 No: 241) 
                   
               
               
                   
               
               
                 11746.0_F 
                 CCCCAAAACCCCAAAACCCCGTATTGATGAT 
                 Primer pair for amplifying 
               
               
                   
                 AAAATTGTATACAAGTTGATAG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 242) 
                 mini-genome 
               
               
                 11746.0_R 
                 CCCCAAAACCCCAAAACCCCTAGATGCTTAA 
                   
               
               
                   
                 TTATTAAGAAGATTCTGGAATG (SEQ ID No: 
                   
               
               
                   
                 243) 
                   
               
               
                   
               
               
                 22291.0_F 
                 CCCCAAAACCCCAAAACCCCATAAACCAATG 
                 Primer pair for amplifying 
               
               
                   
                 TAATTAATTTATTGGGTGTGTTG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 244) 
                 mini-genome 
               
               
                 22291.0_R 
                 CCCCAAAACCCCAAAACCCCTTAGATTAAATT 
                   
               
               
                   
                 TAGAGAGTTATAGAAATGTAGTAAAT (SEQ ID 
                   
               
               
                   
                 No: 245) 
                   
               
               
                   
               
               
                 17535.0_F 
                 CCCCAAAACCCCAAAACCCCATCTCAATTTAT 
                 Primer pair for amplifying 
               
               
                   
                 AAAATCAGAATAAGAGATTGTC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 246) 
                 mini-genome 
               
               
                 17535.0_R 
                 CCCCAAAACCCCAAAACCCCAGAATAAAACA 
                   
               
               
                   
                 ACTGAAGTAAATATGAGTTAC (SEQ ID No: 
                   
               
               
                   
                 247) 
                   
               
               
                   
               
               
                 15372.0_F 
                 CCCCAAAACCCCAAAACCCCTTTCAAATATAA 
                 Primer pair for amplifying 
               
               
                   
                 AATAAACAGAAGAATGGCAAACG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 248) 
                 mini-genome 
               
               
                 15372.0_R 
                 CCCCAAAACCCCAAAACCCCAAATTCAATATT 
                   
               
               
                   
                 AAATGAAATAATTTTCAAAAGTG (SEQ ID 
                   
               
               
                   
                 No: 249) 
                   
               
               
                   
               
               
                 13537.0_F 
                 CCCCAAAACCCCAAAACCCCATGAGATCAAA 
                 Primer pair for amplifying 
               
               
                   
                 TTTTTTTATTAAAATTCTTC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 250) 
                 mini-genome 
               
               
                 13537.0_R 
                 CCCCAAAACCCCAAAACCCCTTGGATTCATA 
                   
               
               
                   
                 TTTTTGTTTAAGGCTTAGATA (SEQ ID No: 
                   
               
               
                   
                 251) 
                   
               
               
                   
               
               
                 22613.0_F 
                 CCCCAAAACCCCAAAACCCCATTAGAAAAGA 
                 Primer pair for amplifying 
               
               
                   
                 GGATTTCAATAAAAGCAAATAT (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 252) 
                 mini-genome 
               
               
                 22613.0_R 
                 CCCCAAAACCCCAAAACCCCATCGATTTATT 
                   
               
               
                   
                 ATTGTTGAATTTAAAAGTATTGAA (SEQ ID 
                   
               
               
                   
                 No: 253) 
                   
               
               
                   
               
               
                 12585.0_F 
                 CCCCAAAACCCCAAAACCCCGAGAGGTTTGA 
                 Primer pair for amplifying 
               
               
                   
                 TAAGTAGAATTAGTAAAATCTATAAAG (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 254) 
                 mini-genome 
               
               
                 12585.0_R 
                 CCCCAAAACCCCAAAACCCCATTAGTACTAT 
                   
               
               
                   
                 TTTCATAGATCTATGTATAAATTGAA (SEQ ID 
                   
               
               
                   
                 No: 255) 
                   
               
               
                   
               
               
                 5317.0_F 
                 CCCCAAAACCCCAAAACCCCAATGGAAAGAT 
                 Primer pair for amplifying 
               
               
                   
                 AAACAGATTTTAATTTGGAAATAAAAT (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 256) 
                 mini-genome 
               
               
                 5317.0_R 
                 CCCCAAAACCCCAAAACCCCTTTAAGCAGTA 
                   
               
               
                   
                 TTTCTAAAATGTTGATGAAATAAAAAT (SEQ 
                   
               
               
                   
                 ID No: 257) 
                   
               
               
                   
               
               
                 17894.0_F 
                 CCCCAAAACCCCAAAACCCCATAAGATAAAA 
                 Primer pair for amplifying 
               
               
                   
                 TTTAACGAAAAAAAGTTAAGTC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 258) 
                 mini-genome 
               
               
                 17894.0_R 
                 CCCCAAAACCCCAAAACCCCATAAGATGAAA 
                   
               
               
                   
                 TATAGAGATAATTGAGCCTA (SEQ ID 
                   
               
               
                   
                 No: 259) 
                   
               
               
                   
               
               
                 3513.0_F 
                 CCCCAAAACCCCAAAACCCCAATTACATATTA 
                 Primer pair for amplifying 
               
               
                   
                 ATGTACTTATGATAGAATG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 260) 
                 mini-genome 
               
               
                 3513.0_R 
                 CCCCAAAACCCCAAAACCCCTAATGATCAAA 
                   
               
               
                   
                 TAACCTGAGTTAAAGAAG (SEQ ID 
                   
               
               
                   
                 No: 261) 
                   
               
               
                   
               
               
                 16420.0_F 
                 CCCCAAAACCCCAAAACCCCAAATTATGAAA 
                 Primer pair for amplifying 
               
               
                   
                 ATAGACACTAATTGGATGTTC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 262) 
                 mini-genome 
               
               
                 16420.0_R 
                 CCCCAAAACCCCAAAACCCCTGATTCGTCAT 
                   
               
               
                   
                 ATGAAATTGAAAAGGAGTAAAT (SEQ ID No: 
                   
               
               
                   
                 263) 
                   
               
               
                   
               
               
                 1084.1_F 
                 CCCCAAAACCCCAAAACCCCAGCGCCATGAA 
                 Primer pair for amplifying 
               
               
                   
                 TCTGATGCATTTATTTTAAG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 264) 
                 mini-genome 
               
               
                 1084.1_R 
                 CCCCAAAACCCCAAAACCCCGTAGATCATTT 
                   
               
               
                   
                 ATGTAAAAGATTTTGAGAGATG (SEQ ID No: 
                   
               
               
                   
                 265) 
                   
               
               
                   
               
               
                 22651.0_F 
                 CCCCAAAACCCCAAAACCCCATACAATTATTA 
                 Primer pair for amplifying 
               
               
                   
                 TAAATGAAAAAGCGCACTAATC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 266) 
                 mini-genome 
               
               
                 22651.0_R 
                 CCCCAAAACCCCAAAACCCCATAGTTACTAT 
                   
               
               
                   
                 GAAAGGACTGGTACATAGAAATAATAG (SEQ 
                   
               
               
                   
                 ID No: 267) 
                   
               
               
                   
               
               
                 8670.0_F 
                 CCCCAAAACCCCAAAACCCCTTAAGTCAATA 
                 Primer pair for amplifying 
               
               
                   
                 TCTAAATCAAATATTAGTAGTATAAT (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 268) 
                 mini-genome 
               
               
                 8670.0_R 
                 CCCCAAAACCCCAAAACCCCGTCATATGGTT 
                   
               
               
                   
                 TTATAAAATAAAATTGAGATTTTTTTG (SEQ 
                   
               
               
                   
                 ID No: 269) 
                   
               
               
                   
               
               
                 19107.0_F 
                 CCCCAAAACCCCAAAACCCCATAAGGATAAA 
                 Primer pair for amplifying 
               
               
                   
                 TTCTATCATATAAGTGGAAGTGC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 270) 
                 mini-genome 
               
               
                 19107.0_R 
                 CCCCAAAACCCCAAAACCCCATTCTTGAATA 
                   
               
               
                   
                 TTGATTATGCATATTGTGTAAAATAG (SEQ ID 
                   
               
               
                   
                 No: 271) 
                   
               
               
                   
               
               
                 21021.0_F 
                 CCCCAAAACCCCAAAACCCCAAGCGTTGAAT 
                 Primer pair for amplifying 
               
               
                   
                 TTTTTATAATATATGATAAAC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 272) 
                 mini-genome 
               
               
                 21021.0_R 
                 CCCCAAAACCCCAAAACCCCTTAATGCCAAT 
                   
               
               
                   
                 AAACAGATGAAAGTAGAGTTATAG (SEQ ID 
                   
               
               
                   
                 No: 273) 
                   
               
               
                   
               
               
                 15004.0_F 
                 CCCCAAAACCCCAAAACCCCATAGAGAGTGT 
                 Primer pair for amplifying 
               
               
                   
                 TTTATTGAAGGACAGAGAATATTG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 274) 
                 mini-genome 
               
               
                 15004.0_R 
                 CCCCAAAACCCCAAAACCCCGAGCGTAAGAA 
                   
               
               
                   
                 ATATTCTTAGATAAATGGAAACTG (SEQ ID 
                   
               
               
                   
                 No: 275) 
                   
               
               
                   
               
               
                 18789.0_F 
                 CCCCAAAACCCCAAAACCCCATGGCAATATC 
                 Primer pair for amplifying 
               
               
                   
                 TTTGCGTGTTTCTGGC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 276) 
                 mini-genome 
               
               
                 18789.0_R 
                 CCCCAAAACCCCAAAACCCCATAAGAATAAA 
                   
               
               
                   
                 TTAAAGAAGATTTGAGAAAGATATGC (SEQ ID 
                   
               
               
                   
                 No: 277) 
                   
               
               
                   
               
               
                 1335.1_F 
                 CCCCAAAACCCCAAAACCCCAAATGCTAAAA 
                 Primer pair for amplifying 
               
               
                   
                 ATAATGAAAAATCTGAGGG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 278) 
                 mini-genome 
               
               
                 1335.1_R 
                 CCCCAAAACCCCAAAACCCCTAATGACAGGT 
                   
               
               
                   
                 TTAGTAATAATTTAGCTG (SEQ ID 
                   
               
               
                   
                 No: 279) 
                   
               
               
                   
               
               
                 17286.0_F 
                 CCCCAAAACCCCAAAACCCCACGACTTAACA 
                 Primer pair for amplifying 
               
               
                   
                 TTGCTGTTAAATATTCAGAAAT (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 280) 
                 mini-genome 
               
               
                 17286.0_R 
                 CCCCAAAACCCCAAAACCCCTAAAATTGGAA 
                   
               
               
                   
                 AGGGGCAAATTTGCTTATGA (SEQ ID No: 
                   
               
               
                   
                 281) 
                   
               
               
                   
               
               
                 7278.0_F 
                 CCCCAAAACCCCAAAACCCCATGAGTAATAT 
                 Primer pair for amplifying 
               
               
                   
                 ATACAAATTTTAAATGTATTTTGATTTA (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 282) 
                 mini-genome 
               
               
                 7278.0_R 
                 CCCCAAAACCCCAAAACCCCATTGAGTGAGT 
                   
               
               
                   
                 ATTTTTATATTTATTGCGAGTTA (SEQ ID 
                   
               
               
                   
                 No: 283) 
                   
               
               
                   
               
               
                 7752.0_F 
                 CCCCAAAACCCCAAAACCCCACAATAGGCAT 
                 Primer pair for amplifying 
               
               
                   
                 ATTTAATAATTAATTGTTAAAG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 284) 
                 mini-genome 
               
               
                 7752.0_R 
                 CCCCAAAACCCCAAAACCCCACTCATTATAT 
                   
               
               
                   
                 AAGGCTGAAAAAATCAGAGG (SEQ ID No: 
                   
               
               
                   
                 285) 
                   
               
               
                   
               
               
                 244.1_F 
                 CCCCAAAACCCCAAAACCCCTAAATGTAAGA 
                 Primer pair for amplifying 
               
               
                   
                 GTAAACTATCATATGAAAG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 286) 
                 mini-genome 
               
               
                 244.1_R 
                 CCCCAAAACCCCAAAACCCCATAATGCGAAA 
                   
               
               
                   
                 TATTCATCAGAGTAAATAATG (SEQ ID No: 
                   
               
               
                   
                 287) 
                   
               
               
                   
               
               
                 20383.0_F 
                 CCCCAAAACCCCAAAACCCCATACGTCATGA 
                 Primer pair for amplifying 
               
               
                   
                 TTATAAGATTATTATAGAATGCTTAC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 288) 
                 mini-genome 
               
               
                 20383.0_R 
                 CCCCAAAACCCCAAAACCCCTCTTGTAAAAT 
                   
               
               
                   
                 AATAAGTTTAAGAAATTGAATTTAG (SEQ ID 
                   
               
               
                   
                 No: 289) 
                   
               
               
                   
               
               
                 331.1_F 
                 CCCCAAAACCCCAAAACCCCATAATATCAAA 
                 Primer pair for amplifying 
               
               
                   
                 TTAATGAATATTTATCAATTTTATTAAT (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 290) 
                 mini-genome 
               
               
                 331.1_R 
                 CCCCAAAACCCCAAAACCCCCCCTAATGTCC 
                   
               
               
                   
                 ATAATTTATGTATCAAATAAGG (SEQ ID No: 
                   
               
               
                   
                 291) 
                   
               
               
                   
               
               
                 22208.0_F 
                 CCCCAAAACCCCAAAACCCCATGATGGTGGA 
                 Primer pair for amplifying 
               
               
                   
                 GGAGTGAAGATAAATTAGAATG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 292) 
                 mini-genome 
               
               
                 22208.0_R 
                 CCCCAAAACCCCAAAACCCCAAAGTGCAATA 
                   
               
               
                   
                 AAAAGAGTGAAAATAAATTTTTG (SEQ ID 
                   
               
               
                   
                 No: 293) 
                   
               
               
                   
               
               
                 21398.0_F 
                 CCCCAAAACCCCAAAACCCCATATACCAATG 
                 Primer pair for amplifying 
               
               
                   
                 TTAAAAATGAATATTGATATAGAATAG (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 294) 
                 mini-genome 
               
               
                 21398.0_R 
                 CCCCAAAACCCCAAAACCCCATAATACAAAG 
                   
               
               
                   
                 TAAAATTGTTTTTTATAGTTCATAA (SEQ ID 
                   
               
               
                   
                 No: 295) 
                   
               
               
                   
               
               
                 11890.0_F 
                 CCCCAAAACCCCAAAACCCCACATAGTGAAT 
                 Primer pair for amplifying 
               
               
                   
                 GAATTAATGAATAAGTTTGAG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 296) 
                 mini-genome 
               
               
                 11890.0_R 
                 CCCCAAAACCCCAAAACCCCGTGATAATAAA 
                   
               
               
                   
                 TTCCTGAGTATATAGTTTAAGAAG (SEQ ID 
                   
               
               
                   
                 No: 297) 
                   
               
               
                   
               
               
                 13521.0_F 
                 CCCCAAAACCCCAAAACCCCGTGATTGCATT 
                 Primer pair for amplifying 
               
               
                   
                 TTTTTGCGAAATATTTGC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 298) 
                 mini-genome 
               
               
                 13521.0_R 
                 CCCCAAAACCCCAAAACCCCTGAGTTCTCAT 
                   
               
               
                   
                 GTAATAAAAGAATCCATG (SEQ ID No: 
                   
               
               
                   
                 299) 
                   
               
               
                   
               
               
                 3511.0_F 
                 CCCCAAAACCCCAAAACCCCATGATGCTACA 
                 Primer pair for amplifying 
               
               
                   
                 AAAACGCTATATAATCTATAAC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 300) 
                 mini-genome 
               
               
                 3511.0_R 
                 CCCCAAAACCCCAAAACCCCTTGAACTTTCA 
                   
               
               
                   
                 ATAGATGTTTGATTAAATTC (SEQ ID 
                   
               
               
                   
                 No: 301) 
                   
               
               
                   
               
               
                 22209.0_F 
                 CCCCAAAACCCCAAAACCCCAAAGATATGTG 
                 Primer pair for amplifying 
               
               
                   
                 GCTGGATTTTAAAATATGGTTG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 302) 
                 mini-genome 
               
               
                 22209.0_R 
                 CCCCAAAACCCCAAAACCCCAAGACTAATGA 
                   
               
               
                   
                 ATTTGAGAATTATAAAATAATGAATC (SEQ ID 
                   
               
               
                   
                 No: 303) 
                   
               
               
                   
               
               
                 18924.0_F 
                 CCCCAAAACCCCAAAACCCCATCAACTTTAA 
                 Primer pair for amplifying 
               
               
                   
                 TTCATTGTAGGAATTAAAGATGTAATAC (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 304) 
                 mini-genome 
               
               
                 18924.0_R 
                 CCCCAAAACCCCAAAACCCCGTGAGAACAAA 
                   
               
               
                   
                 TAATAATAAAAATAAAGGAATTAA (SEQ ID 
                   
               
               
                   
                 No: 305) 
                   
               
               
                   
               
               
                 14977.0_F 
                 CCCCAAAACCCCAAAACCCCAATTCTTTATCT 
                 Primer pair for amplifying 
               
               
                   
                 GAATTAGATAAGAATTCATAAGC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 306) 
                 mini-genome 
               
               
                 14977.0_R 
                 CCCCAAAACCCCAAAACCCCGTGAGTATGCA 
                   
               
               
                   
                 ATAGATTGTTAATTAAATTTG (SEQ ID No: 
                   
               
               
                   
                 307) 
                   
               
               
                   
               
               
                 18694.0_F 
                 CCCCAAAACCCCAAAACCCCAAGTTGCTAAA 
                 Primer pair for amplifying 
               
               
                   
                 AATAGTTGATAGCAACAAGTTAT (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 308) 
                 mini-genome 
               
               
                 18694.0_R 
                 CCCCAAAACCCCAAAACCCCTGGATGTGTTT 
                   
               
               
                   
                 TTTTCCAAATTAATGAACAAAAATTAAA (SEQ 
                   
               
               
                   
                 ID No: 309) 
                   
               
               
                   
               
               
                 13237.0_F 
                 CCCCAAAACCCCAAAACCCCAACATTCTAAA 
                 Primer pair for amplifying 
               
               
                   
                 TTTCTTCTTTATAAGATTATTG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 310) 
                 mini-genome 
               
               
                 13237.0_R 
                 CCCCAAAACCCCAAAACCCCATCTAAACTAA 
                   
               
               
                   
                 TCTGAAACCAAAGATAGTATG (SEQ ID No: 
                   
               
               
                   
                 311) 
                   
               
               
                   
               
               
                 21338.0_F 
                 CCCCAAAACCCCAAAACCCCGTTATCCATAT 
                 Primer pair for amplifying 
               
               
                   
                 ATACGTAAGCATTTTGCGATTG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 312) 
                 mini-genome 
               
               
                 21338.0_R 
                 CCCCAAAACCCCAAAACCCCGAAACCTATGC 
                   
               
               
                   
                 ATTATTTTTAAAGAAATATTAAATTAA (SEQ 
                   
               
               
                   
                 ID No: 313) 
                   
               
               
                   
               
               
                 215.1_F 
                 CCCCAAAACCCCAAAACCCCTCGTACATTAA 
                 Primer pair for amplifying 
               
               
                   
                 TAGTTGAAATTGCTTTTATTAAATTG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 314) 
                 mini-genome 
               
               
                 215.1_R 
                 CCCCAAAACCCCAAAACCCCGTAGTCTAAAA 
                   
               
               
                   
                 TAAATTTTATTTTGGGTTTTAA (SEQ ID No: 
                   
               
               
                   
                 315) 
                   
               
               
                   
               
               
                 13236.0_F 
                 CCCCAAAACCCCAAAACCCCGTTAAATGATA 
                 Primer pair for amplifying 
               
               
                   
                 ATCATAGCAAAATTGCGGTAT (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 316) 
                 mini-genome 
               
               
                 13236.0_R 
                 CCCCAAAACCCCAAAACCCCAAGGATAAATA 
                   
               
               
                   
                 TTGAAAGTAAATGTTCTAATTAATTTGC (SEQ 
                   
               
               
                   
                 ID No: 317) 
                   
               
               
                   
               
               
                 16827.0_F 
                 CCCCAAAACCCCAAAACCCCAGAAATGAAAA 
                 Primer pair for amplifying 
               
               
                   
                 GAATGATTTTTGAGGGGATTC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 318) 
                 mini-genome 
               
               
                 16827.0_R 
                 CCCCAAAACCCCAAAACCCCTAAAGGCAAAA 
                   
               
               
                   
                 GTCGATTTAAATGCTCAGTTTC (SEQ ID No: 
                   
               
               
                   
                 319) 
                   
               
               
                   
               
               
                 15136.0_F 
                 CCCCAAAACCCCAAAACCCCTTAAGGCTAAA 
                 Primer pair for amplifying 
               
               
                   
                 ATACTTGTTTTACTAGAGAAC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 320) 
                 mini-genome 
               
               
                 15136.0_R 
                 CCCCAAAACCCCAAAACCCCATAAATCAAAT 
                   
               
               
                   
                 TAAATTGCATAACATGAAC (SEQ ID 
                   
               
               
                   
                 No: 321) 
                   
               
               
                   
               
               
                 115.1_F 
                 CCCCAAAACCCCAAAACCCCAGAGGATGTAA 
                 Primer pair for amplifying 
               
               
                   
                 ATTACAATAAATCGTAAAAAC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 322) 
                 mini-genome 
               
               
                 115.1_R 
                 CCCCAAAACCCCAAAACCCCTTCTAAAAAAT 
                   
               
               
                   
                 ATAAAGATAAATTGACGTC (SEQ ID 
                   
               
               
                   
                 No: 323) 
                   
               
               
                   
               
               
                 21295.0_F 
                 CCCCAAAACCCCAAAACCCCATCCAGTTGAA 
                 Primer pair for amplifying 
               
               
                   
                 ATCTAAAACAATTTTGTATATTTAAAG (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 324) 
                 mini-genome 
               
               
                 21295.0_R 
                 CCCCAAAACCCCAAAACCCCTTAAGAGATTG 
                   
               
               
                   
                 CATTATAAATAAGATAGGATTC (SEQ ID No: 
                   
               
               
                   
                 325) 
                   
               
               
                   
               
               
                 16269.0_F 
                 CCCCAAAACCCCAAAACCCCATTGATTGATA 
                 Primer pair for amplifying 
               
               
                   
                 AACTTGGAAGTTAAGAAAGATTTG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 326) 
                 mini-genome 
               
               
                 16269.0_R 
                 CCCCAAAACCCCAAAACCCCATGAATAACAG 
                   
               
               
                   
                 ATGGAATGCTTCAAGATATG (SEQ ID No: 
                   
               
               
                   
                 327) 
                   
               
               
                   
               
               
                 644.1_F 
                 CCCCAAAACCCCAAAACCCCAAATGTTAGTA 
                 Primer pair for amplifying 
               
               
                   
                 TTTGAATTAAAGAGAGGTAAAAC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 328) 
                 mini-genome 
               
               
                 644.1_R 
                 CCCCAAAACCCCAAAACCCCTTATGAAAATG 
                   
               
               
                   
                 AAATGGTTTTGATTGGCTAATAA (SEQ ID 
                   
               
               
                   
                 No: 329) 
                   
               
               
                   
               
               
                 5586.0_F 
                 CCCCAAAACCCCAAAACCCCATGAGTAAAAT 
                 Primer pair for amplifying 
               
               
                   
                 TTAGCTTAAGTAATGTAAGAATC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 330) 
                 mini-genome 
               
               
                 5586.0_R 
                 CCCCAAAACCCCAAAACCCCATATATCAAAA 
                   
               
               
                   
                 TATCAACATTTTTTTGTGTGATTGTTAC (SEQ 
                   
               
               
                   
                 ID No: 331) 
                   
               
               
                   
               
               
                 13085.0_F 
                 CCCCAAAACCCCAAAACCCCTTGATGAAATT 
                 Primer pair for amplifying 
               
               
                   
                 TGAAAATGAATAGAGAGTAC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 332) 
                 mini-genome 
               
               
                 13085.0_R 
                 CCCCAAAACCCCAAAACCCCGTAATGCTACA 
                   
               
               
                   
                 TTTGCAAAAAAGTACAAACAG (SEQ ID No: 
                   
               
               
                   
                 333) 
                   
               
               
                   
               
               
                 13838.0_F 
                 CCCCAAAACCCCAAAACCCCGTAAGGCCAGA 
                 Primer pair for amplifying 
               
               
                   
                 ATCAATGAATAAAAAGGTC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 334) 
                 mini-genome 
               
               
                 13838.0_R 
                 CCCCAAAACCCCAAAACCCCGAAAAGGGAG 
                   
               
               
                   
                 ATTTACAAAAATTTGTAGATGTTATATTG 
                   
               
               
                   
                 (SEQ ID No: 335) 
                   
               
               
                   
               
               
                 1415.1_F 
                 CCCCAAAACCCCAAAACCCCATTGATCATTA 
                 Primer pair for amplifying 
               
               
                   
                 ATAAAGAAGAATTGCTAATAT (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 336) 
                 mini-genome 
               
               
                 1415.1_R 
                 CCCCAAAACCCCAAAACCCCAATGCGATGAA 
                   
               
               
                   
                 ATGTTTTTTATTATGAAAAG (SEQ ID 
                   
               
               
                   
                 No: 337) 
                   
               
               
                   
               
               
                 19468.0_F 
                 CCCCAAAACCCCAAAACCCCAAGGAAGTTCA 
                 Primer pair for amplifying 
               
               
                   
                 ATGCTATTTAGCAAATTAGG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 338) 
                 mini-genome 
               
               
                 19468.0_R 
                 CCCCAAAACCCCAAAACCCCTTGATTCAAAA 
                   
               
               
                   
                 TATGCACAAGATTAAAAATTCAC (SEQ ID 
                   
               
               
                   
                 No: 339) 
                   
               
               
                   
               
               
                 20407.0_F 
                 CCCCAAAACCCCAAAACCCCATAAGAAAGAT 
                 Primer pair for amplifying 
               
               
                   
                 AAGTTGCAATTAAATAATAAGG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 340) 
                 mini-genome 
               
               
                 20407.0_R 
                 CCCCAAAACCCCAAAACCCCATGAAGACAAG 
                   
               
               
                   
                 TCTGATGAAAATAGAATGG (SEQ ID No: 
                   
               
               
                   
                 341) 
                   
               
               
                   
               
               
                 19922.0_F 
                 CCCCAAAACCCCAAAACCCCATAGTCTTAAA 
                 Primer pair for amplifying 
               
               
                   
                 ATTTTATACTATCATGAAATAATATTAAG (SEQ 
                 chromosome, to be added to 
               
               
                   
                 ID No: 342) 
                 mini-genome 
               
               
                 19922.0_R 
                 CCCCAAAACCCCAAAACCCCGTAAGTCTAAA 
                   
               
               
                   
                 GTTTAACAGTTTTTAGTAAATATC (SEQ ID 
                   
               
               
                   
                 No: 343) 
                   
               
               
                   
               
               
                 20459.0_F 
                 CCCCAAAACCCCAAAACCCCTTATGCTAGTT 
                 Primer pair for amplifying 
               
               
                   
                 GAGTGATTGAAAATATATTTGTGC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 344) 
                 mini-genome 
               
               
                 20459.0_R 
                 CCCCAAAACCCCAAAACCCCTTGACGTAGAA 
                   
               
               
                   
                 TAATGGGCTTATAGAAG (SEQ ID No: 
                   
               
               
                   
                 345) 
                   
               
               
                   
               
               
                 20493.0_F 
                 CCCCAAAACCCCAAAACCCCTTAATCAACTC 
                 Primer pair for amplifying 
               
               
                   
                 ACTTTACCCACTAATCAAACAC (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 346) 
                 mini-genome 
               
               
                 20493.0_R 
                 CCCCAAAACCCCAAAACCCCATATTTAAGAT 
                   
               
               
                   
                 ATACAGAAATATAGAGAATACAAC (SEQ ID 
                   
               
               
                   
                 No: 347) 
                   
               
               
                   
               
               
                 9925.0_F 
                 CCCCAAAACCCCAAAACCCCATTGGATCAAT 
                 Primer pair for amplifying 
               
               
                   
                 TTTGAAGAGAATTCATGGAAAAT (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 348) 
                 mini-genome 
               
               
                 9925.0_R 
                 CCCCAAAACCCCAAAACCCCATCAGAAAAAA 
                   
               
               
                   
                 TATTTGAAAATTCGATAAAGC (SEQ ID No: 
                   
               
               
                   
                 349) 
                   
               
               
                   
               
               
                 22456.0_F 
                 CCCCAAAACCCCAAAACCCCATTTCACTTTAT 
                 Primer pair for amplifying 
               
               
                   
                 TTATATATAGATTTGAAATTAAAGTT (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 350) 
                 mini-genome 
               
               
                 22456.0_R 
                 CCCCAAAACCCCAAAACCCCAGTTGACATGT 
                   
               
               
                   
                 TATTTCCAAATTTTCATGGATA (SEQ ID No: 
                   
               
               
                   
                 351) 
                   
               
               
                   
               
               
                 17712.0_F 
                 CCCCAAAACCCCAAAACCCCATGATAACAGG 
                 Primer pair for amplifying 
               
               
                   
                 AATATTTTATAAAATAGTTAAG (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 352) 
                 mini-genome 
               
               
                 17712.0_R 
                 CCCCAAAACCCCAAAACCCCTCACTCTATGC 
                   
               
               
                   
                 AATAAATTTGTTGATATATT (SEQ ID No: 
                   
               
               
                   
                 353) 
                   
               
               
                   
               
               
                 11116.0_F 
                 CCCCAAAACCCCAAAACCCCTTAAAAAAAGA 
                 Primer pair for amplifying 
               
               
                   
                 ATAGTTGGAATAAAAATGAATTT (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 354) 
                 mini-genome 
               
               
                 11116.0_R 
                 CCCCAAAACCCCAAAACCCCAATAGATAAAG 
                   
               
               
                   
                 ATGCCTTTTTTAATAAGTATTTAAC (SEQ ID 
                   
               
               
                   
                 No: 355) 
                   
               
               
                   
               
               
                 19275.0_F 
                 CCCCAAAACCCCAAAACCCCGAGAGGATAAA 
                 Primer pair for amplifying 
               
               
                   
                 TTTATATGAAAATAAAAATAAAGC (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 356) 
                 mini-genome 
               
               
                 19275.0_R 
                 CCCCAAAACCCCAAAACCCCATAAATAAGAA 
                   
               
               
                   
                 ATTTTAAGAATAACGGGCAAATTAG (SEQ ID 
                   
               
               
                   
                 No: 357) 
                   
               
               
                   
               
               
                 21217.0_F 
                 CCCCAAAACCCCAAAACCCCTTGAATTTTAAA 
                 Primer pair for amplifying 
               
               
                   
                 TAAACTTCTTTGTATGATTTAAATG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 358) 
                 mini-genome 
               
               
                 21217.0_R 
                 CCCCAAAACCCCAAAACCCCATAGATTACTT 
                   
               
               
                   
                 TTCAAAGAATTTCTTGACATTC (SEQ ID No: 
                   
               
               
                   
                 359) 
                   
               
               
                   
               
               
                 10537.0_F 
                 CCCCAAAACCCCAAAACCCCAAAGCAAAGAA 
                 Primer pair for amplifying 
               
               
                   
                 ATCTGATGTTTTATTAGAAAAAGTG (SEQ ID 
                 chromosome, to be added to 
               
               
                   
                 No: 360) 
                 mini-genome 
               
               
                 10537.0_R 
                 CCCCAAAACCCCAAAACCCCATGAGATGATA 
                   
               
               
                   
                 ATATTGCCTTTTTGCATATAAT (SEQ ID No: 
                   
               
               
                   
                 361) 
                   
               
               
                   
               
               
                 22670.0_F 
                 CCCCAAAACCCCAAAACCCCATCCTTATACA 
                 Primer pair for amplifying 
               
               
                   
                 AATTCAGAAAACTTAGCAAAT (SEQ ID No: 
                 chromosome, to be added to 
               
               
                   
                 362) 
                 mini-genome 
               
               
                 22670.0_R 
                 CCCCAAAACCCCAAAACCCCGTGGAGAATTT 
                   
               
               
                   
                 TCTAAAGAATTTTCGGAAATTTG (SEQ ID 
                   
               
               
                   
                 No: 363) 
                   
               
               
                   
               
               
                 1781.0_F 
                 CCCCAAAACCCCAAAACCCCACTAGTCTTAA 
                 PCR primers for amplifying 
               
               
                   
                 ATATGAGAAAGATGATTTGAATAAG (SEQ ID 
                 synthetic chromosome 1 and 6 
               
               
                   
                 No: 364) 
                 in FIG. 5B 
               
               
                 1781.0_R 
                 CCCCAAAACCCCAAAACCCCACAAGATAACA 
                   
               
               
                   
                 TTGCTAATTTTCAATAAATTAAAT (SEQ ID 
                   
               
               
                   
                 No: 365) 
                   
               
               
                   
               
               
                 1781.0_Purple_F 
                 GTCAGTGGTCTCAGTATGAAATTTACCTTGG 
                 PCR primers for amplifying 
               
               
                   
                 ATCCTCAGTGTTTCCCTTTGTG (SEQ ID No: 
                 purple DNA building block in 
               
               
                   
                 366) 
                 synthetic chromosomes 2-4 in 
               
               
                 1781.0_Purple_R 
                 AACGCTCGGTCTCGCAGAAATAAATAATAGG 
                 FIG. 5B 
               
               
                   
                 CAGCATAATAATTCAGG (SEQ ID No: 
                   
               
               
                   
                 367) 
                   
               
               
                   
               
               
                 1781.0_red_F 
                 GTCAGTGGTCTCTCCAGTGGATTCACATCAC 
                 PCR primers for amplifying 
               
               
                   
                 TACCATTG (SEQ ID No: 368) 
                 red DNA building block in 
               
               
                 1781.0_red_R 
                 CCCCAAAACCCCAAAACCCCACAAGATAACA 
                 synthetic chromosomes 2-4 
               
               
                   
                 TTGCTAATTTTCAATAAATTAAAT (SEQ ID 
                 in FIG. 5B 
               
               
                   
                 No: 369) 
                   
               
               
                   
               
               
                 1781.0_turquoise_F 
                 CCCCAAAACCCCAAAACCCCACTAGTCTTAA 
                 PCR primers for amplifying 
               
               
                   
                 ATATGAGAAAGATGATTTGAATAAG (SEQ ID 
                 turquoise DNA building block 
               
               
                   
                 No: 370) 
                 in synthetic chromosomes 2-4 
               
               
                 1781.0_turquoise_R 
                 ACGCTCGGTCTCGATACAATCATTAGCATAT 
                 in FIG. 5B 
               
               
                   
                 ACATGCAGTCTGCTTAAATTAAG (SEQ ID 
                   
               
               
                   
                 No: 371) 
                   
               
               
                   
               
               
                 DarkBlue_6mA_top 
                 TCTGTAATTAATGGATCACTATAATCATCTGG 
                 Oligos for annealing to make 
               
               
                   
                 ATGACTATTGGTATTTGATGACGGTGCACAT 
                 blue DNA building block in 
               
               
                   
                 TTGACTTCTT (SEQ ID No: 372) 
                 synthetic chromosomes 2-4 in 
               
               
                 DarkBlue_6mA_bottom 
                 ATTAATTACCTAGTGATATTAGTAGACCTACT 
                 FIG. 5B. Bold red nucleotides 
               
               
                   
                 GATAACCATAAACTACTGCCACGTGTAAACT 
                 represent 6mA. 
               
               
                   
                 GAAGAAGGTC (SEQ ID No: 373) 
                   
               
               
                   
               
               
                 1781.0_red2_F 
                 AGCCTAGGTCTCGTTCTTTTTGAGGGCGAGG 
                 PCR primers for amplifying 
               
               
                   
                 TTATTGTTAAT (SEQ ID No: 374) 
                 red DNA building block in 
               
               
                 1781.0_red2_R 
                 CCCCAAAACCCCAAAACCCCACAAGATAACA 
                 synthetic chromosome 5 in 
               
               
                   
                 T (SEQ ID No: 375) 
                 FIG. 5B 
               
               
                   
               
               
                 1781.0_orange_F 
                 TAGTCAGGTCTCTAGAATAGGCTCACTCTAA 
                 PCR primers for amplifying 
               
               
                   
                 ATTCGAGTGCAAT (SEQ ID No: 376) 
                 orange DNA building block in 
               
               
                 1781.0_orange_R 
                 TCTACTGGTCTCAGTATGAAATTTACCTTGGA 
                 synthetic chromosome 5 in FIG. 
               
               
                   
                 TCCTCAGTGT (SEQ ID No: 377) 
                 5B 
               
               
                   
               
               
                 1781.0_emerald_F 
                 ATCGTAGGTCTCAATACAATCATTAGCATATA 
                 PCR primers for amplifying 
               
               
                   
                 CATGCAGT (SEQ ID No: 378) 
                 emerald DNA building block in 
               
               
                 1781.0_emerald_R 
                 CCCCAAAACCCCAAAACCCCACTAGTCTTAA 
                 synthetic chromosome 5 in FIG. 
               
               
                   
                 AT (SEQ ID No: 379) 
                 5B 
               
               
                   
               
               
                 17535.0_F 
                 CCCCAAAACCCCAAAACCCCATCTCAATTTAT 
                 PCR primers for amplifying 
               
               
                   
                 AAAATCAGAATAAGAGATTGTC (SEQ ID No: 
                 “buffer” chromosome 
               
               
                   
                 380) 
                 (Contig17535.0) for use in 
               
               
                 17535.0_R 
                 CCCCAAAACCCCAAAACCCCAGAATAAAACA 
                 chromatin assembly 
               
               
                   
                 ACTGAAGTAAATATGAGTTAC (SEQ ID No: 
                   
               
               
                   
                 381) 
                   
               
               
                   
               
               
                 12701assay_F 
                 AAGAAGAACTAGCCAGCTCTCACTCAGTTC 
                 PCR primers for assaying the 
               
               
                   
                 (SEQ ID No: 382) 
                 presence of ectopic DNA 
               
               
                 12701assay_R 
                 TGTCTATCTCATCAGGCTCATCAGCATAGG 
                 insertion in mta1 mutants 
               
               
                   
                 (SEQ ID No: 383) 
                   
               
               
                   
               
               
                 12701_firstround_T7_F 
                 AAGAAGAACTAGCCAGCTCTCACTCAGTTC 
                 PCR primers to generate DNA 
               
               
                   
                 (SEQ ID No: 384) 
                 template for ssRNA in vitro 
               
               
                 12701_firstround_T7_R 
                 CCTCTCTGCCCACTAAATTATTCTGACAGC 
                 transcription. This ssRNA is 
               
               
                   
                 (SEQ ID No: 385) 
                 injected into Oxytricha cells to 
               
               
                   
                   
                 induce ectopic DNA retention 
               
               
                   
                   
                 in MTA1 gene. PCR product is 
               
               
                   
                   
                 amplified from Oxytricha 
               
               
                   
                   
                 gDNA of cell strain JRB310. 
               
               
                   
                   
                 The resulting PCR product is 
               
               
                   
                   
                 subjected to a second round 
               
               
                   
                   
                 of PCR amplification using 
               
               
                   
                   
                 primers 
               
               
                   
                   
                 “12701_secondround_T7_F” 
               
               
                   
                   
                 and 
               
               
                   
                   
                 “12701_secondround_T7_R”. 
               
               
                   
                   
                 The final, second round PCR 
               
               
                   
                   
                 product is then used for 
               
               
                   
                   
                 ssRNA in vitro transcription. 
               
               
                   
               
               
                 12701_secondround_T7_F 
                 CTACTTGATATAATACGACTCACTATAGGGAA 
                 PCR primers for second round 
               
               
                   
                 TTCCTAAGGGGAGTGAAGCCAACAACAG 
                 amplification of DNA template, 
               
               
                   
                 (SEQ ID No: 386) 
                 to be used for ssRNA in vitro 
               
               
                 12701_secondround_T7_R 
                 TGTCTATCTCATCAGGCTCATCAGCATAGG 
                 transcription. Forward primer 
               
               
                   
                 (SEQ ID No: 387) 
                 contains T7 promoter 
               
               
                   
                   
                 sequence, which is required 
               
               
                   
                   
                 for subsequent in vitro 
               
               
                   
                   
                 transcription. 
               
               
                   
               
               
                 metGATC_F2 
                 GTGCTATGCATTTTAAATTTATTCGCATTGAA 
                 PCR primers for amplification 
               
               
                   
                 GA (SEQ ID No: 388) 
                 of DNA substrate for use in 
               
               
                 metGATC_R2 
                 ATTCAGAATTTTAGTGTGTGGAGTATGATAGT 
                 6mA methyltransferase assay 
               
               
                   
                 A (SEQ ID No: 389) 
                 involving Tetrahymena nuclear 
               
               
                   
               
               
                 noGATC2_F 
                 GGTCTATATTATTTTAGTATTCTTTCTATAAAT 
                 PCR primers for amplifying 
               
               
                   
                 G (SEQ ID No: 390) 
                 350 bp dsDNA substrate for 
               
               
                 noGATC2_R 
                 GTTACAAGAATATAAGAAAAGAAAGGGTGAA 
                 methyltransferase assays 
               
               
                   
                 TAGG (SEQ ID No: 391) 
                 involving recombinant proteins 
               
               
                   
                   
                 (in FIGS. 2E, 2F, and 10H) 
               
               
                   
               
               
                 T7noGATC2_F2 
                 TAATACGACTCACTATAGGG 
                 PCR primers for amplifying 
               
               
                   
                 GGTCTATATTATTTTAGTATTCTTTC (SEQ ID 
                 DNA ~350 bp dsDNA template 
               
               
                   
                 No: 392) 
                 with T7 overhangs at one end, 
               
               
                 noGATC2_R 
                 GTTACAAGAATATAAGAAAAGAAAGGGTGAA 
                 for subsequent ssRNA 
               
               
                   
                 TAGG (SEQ ID No: 393) 
                 production by in vitro 
               
               
                   
                   
                 transcription 
               
               
                   
               
               
                 T7noGATC2_F2 
                 TAATACGACTCACTATAGGG 
                 PCR primers for amplifying 
               
               
                   
                 GGTCTATATTATTTTAGTATTCTTTC (SEQ ID 
                 DNA ~350 bp dsDNA template 
               
               
                   
                 No: 394) 
                 with T7 overhangs at the 5′ 
               
               
                 T7noGATC2_R2 
                 TAATACGACTCACTATAGGG 
                 and 3′ ends, for subsequent 
               
               
                   
                 GTTACAAGAATATAAGAAAAG (SEQ ID No: 
                 dsRNA production by in vitro 
               
               
                   
                 395) 
                 transcription 
               
               
                   
               
               
                 noGATC_F3 
                 AACTTCTGTCATTACATTAAGCTTTAA (SEQ 
                 DNA oligonucleotides for use 
               
               
                   
                 ID No: 396) 
                 in DNA methyltransferase 
               
               
                 noGATC_R3 
                 TTAAAGCTTAATGTAATGACAGAAGTT (SEQ 
                 assays in FIGS. 2G, 10I, 
               
               
                   
                 ID No: 397) 
                 10J, 10K, 10L 
               
               
                 noGATC_F12 
                 AACTTCTGTCATTAACTTAAGCTTTAA (SEQ 
                   
               
               
                   
                 ID No: 398) 
                   
               
               
                 noGATC_R12 
                 TTAAAGCTTAAGTTAATGACAGAAGTT (SEQ 
                   
               
               
                   
                 ID No: 399) 
                   
               
               
                 noGATC_F13 
                 AACTTCTGTACTTACATTAAGCTTTAA (SEQ 
                   
               
               
                   
                 ID No: 400) 
                   
               
               
                 noGATC_R13 
                 TTAAAGCTTAATGTAAGTACAGAAGTT (SEQ 
                   
               
               
                   
                 ID No: 401) 
                   
               
               
                 noGATC_F14 
                 AACTTCTGTACTTAACTTAAGCTTTAA (SEQ 
                   
               
               
                   
                 ID No: 402) 
                   
               
               
                 noGATC_R14 
                 TTAAAGCTTAAGTTAAGTACAGAAGTT (SEQ 
                   
               
               
                   
                 ID No: 403) 
                   
               
               
                 noGATC_F1 
                 AACTTCTGTCATTACATTAAGCTTTAAAAAAT 
                   
               
               
                   
                 TCAATTCCTTTTATT (SEQ ID No: 404) 
                   
               
               
                 noGATC_R1 
                 AATAAAAGGAATTGAATTTTTTAAAGCTTAAT 
                   
               
               
                   
                 GTAATGACAGAAGTT (SEQ ID No: 405) 
                   
               
               
                 noGATC_F2 
                 TGTCATTACATTAAGCTTTAAAAAATTCAATT 
                   
               
               
                   
                 CCT (SEQ ID No: 406) 
                   
               
               
                 noGATC_R2 
                 AGGAATTGAATTTTTTAAAGCTTAATGTAATG 
                   
               
               
                   
                 ACA (SEQ ID No: 407) 
                   
               
               
                 noGATC_F3 
                 AACTTCTGTCATTACATTAAGCTTTAA (SEQ 
                   
               
               
                   
                 ID No: 408) 
                   
               
               
                 noGATC_R3 
                 TTAAAGCTTAATGTAATGACAGAAGTT (SEQ 
                   
               
               
                   
                 ID No: 409) 
                   
               
               
                 noGATC_F8 
                 TATTAGAATTATGTTCTTCATGAAATT (SEQ 
                   
               
               
                   
                 ID No: 410) 
                   
               
               
                 noGATC_R8 
                 AATTTCATGAAGAACATAATTCTAATA (SEQ 
                   
               
               
                   
                 ID No: 411) 
               
               
                   
               
            
           
         
       
     
                     TABLE 3               Recombinant protein sequences.                                    &gt;MTA1 (manually curated from Tetrahymena DB gene       ID: TTHERM_00704040)       (SEQ ID No: 412)       MSKAVNKKGLRPRKSDSILDHIKNKLDQEFLEDNENGEQSDEDYDQKS               LNKAKKPYKKRQTQNGSELVISQQKTKAKASANNKKSAKNSQKLDEEE               KIVEEEDLSPQKNGAVSEDDQQQEASTQEDDYLDRLPKSKKGLQGLLQ               DIEKRILHYKQLFFKEQNEIANGKRSMVPDNSIPICSDVTKLNFQALI               DAQMRHAGKMFDVIMMDPPWQLSSSQPSRGVAIAYDSLSDEKIQNMPI               QSLQQDGFIFVWAINAKYRVTIKMIENWGYKLVDEITWVKKTVNGKIA               KGHGFYLQHAKESCLIGVKGDVDNGRFKKNIASDVIFSERRGQSQKPE               EIYQYINQLCPNGNYLEIFARRNNLHDNWVSIGNEL               &gt;MTA9 (manually curated from Tetrahymena DB gene       ID: TTHERM_00301770)       (SEQ ID No: 413)       MAPKKQEQEPIRLSTRTASKKVDYLQLSNGKLEDFFDDLEEDNKPARN               RSRSKKRGRKPLKKADSRSKTPSRVSNARGRSKSLGPRKTYPRKKNLS               PDNQLSLLLKWRNDKIPLKSASETDNKCKVVNVKNIFKSDLSKYGANL               QALFINALWKVKSRKEKEGLNINDLSNLKIPLSLMKNGILFIWSEKEI               LGQIVEIMEQKGFTYIENFSIMFLGLNKCLQSINHKDEDSQNSTASTN               NTNNEAITSDLTLKDTSKFSDQIQDNHSEDSDQARKQQTPDDITQKKN               KLLKKSSVPSIQKLFEEDPVQTPSVNKPIEKSIEQVTQEKKFVMNNLD               ILKSTDINNLFLRNNYPYFKKTRHTLLMFRRIGDKNQKLELRHQRTSD               VVFEVTDEQDPSKVDTMMKEYVYQMIETLLPKAQFIPGVDKHLKMMEL               FASTDNYRPGWISVIEK               &gt;p1 (manually curated from Tetrahymena DB gene       ID: TTHERM_00161750)       (SEQ ID No: 414)       MSLKKGKFQHNQSKSLWNYTLSPGWREEEVKILKSALQLFGIGKWKKI               MESGCLPGKSIGQIYMQTQRLLGQQSLGDFMGLQIDLEAVFNQNMKKQ               DVLRKNNCIINTGDNPTKEERKRRIEQNRKIYGLSAKQIAEKLPKVKK               HAPQYMTLEDIENEKFTNLEILTHLYNLKAEIVRRLAEQGETIAQPSI               IKSLNNLNHNLEQNQNSNSSTETKVTLEQSGKKKYKVLAIEETELQNG               PIATNSQKKSINGKRKNNRKINSDSEGNEEDISLEDIDSQESEINSEE               IVEDDEEDEQIEEPSKIKKRKKNPEQESEEDDIEEDQEEDELVVNEEE               IFEDDDDDEDNQDSSEDDDDDED               &gt;p2 (manually curated from Tetrahymena DB gene       ID: TTHERM_00439330)       (SEQ ID No: 415)       MKKNSKSQNQPLDFTQYAKNMRKDLSNQDICLEDGALNHSYFLTKKGQ               YWTPLNQKALQRGIELFGVGNWKEINYDEFSGKANIVELELRICMILG               INDITEYYGKKISEEEQEEIKKSNIAKGKKENKLKDNIYQKLQQMQ                    
Sequences were manually curated by mapping RNaseq reads to reference gene annotations and verifying the accuracy of predicted exon boundaries.
 
     Example 2 
     Epigenomic Profiles of Chromatin and Transcription in  Oxytricha    
     We generated genome-wide in vivo maps of nucleosome positioning, transcription, and 6 mA in the macronuclei of asexually growing (vegetative)  Oxytricha trifallax  cells using Mnase sequencing (MNase-seq), poly(Ar RNA sequencing (RNA-seq), transcriptional start site sequencing (TSS-seq), and single-molecule real-time sequencing (SMRT-seq) ( FIGS. 1A-1E ). The smallest  Oxytricha  chromosome is only 430 bp in length, with a single well-positioned nucleosome. Strikingly, 6 mA is enriched in three consecutive nucleosome-depleted regions directly downstream of transcription start sites (TSSs;  FIG. 1A ). Each region contains varying levels of 6 mA ( FIG. 1B ), with the +1/+2 nucleosome linker being most densely methylated (Table 4). In general, highly transcribed chromosomes tend to bear more 6 mA, suggesting a positive role of this DNA modification in gene regulation ( FIG. 1C ). The majority of methylation marks are located within an ApT motif ( FIGS. 1D and 1E ). 6 mA occurs on sense and antisense strands with approximately equal frequency, indicating that the underlying methylation machinery does not function strand-specifically. Quantitative LC-MS/MS analysis confirmed the presence of 6 mA in  Oxytricha  ( FIGS. 8A and 8B ; see Example 1). 
     
       
         
           
               
             
               
                 TABLE 4 
               
             
            
               
                   
               
               
                 Descriptive statistics of 6mA distribution in the genome. 
               
            
           
           
               
               
            
               
                   
                 Number of 6mA sites 
               
            
           
           
               
               
               
            
               
                   
                 Oxytricha 
                 Tetrahymena 
               
            
           
           
               
               
               
               
               
               
               
               
               
               
               
            
               
                   
                   
                   
                   
                   
                 Standard 
                   
                   
                   
                   
                 Standard 
               
               
                   
                 Minimum 
                 Maximum 
                 Median 
                 Mean 
                 Deviation 
                 Minimum 
                 Maximum 
                 Median 
                 Mean 
                 Deviation 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
               
               
            
               
                 Methyl 
                 0 
                 14 
                 2 
                 2.03 
                 2.27 
                 0 
                 27 
                 10 
                 9.66 
                 6.10 
               
               
                 Cluster 1 
               
               
                 Methyl 
                 0 
                 24 
                 6 
                 5.99 
                 4.24 
                 0 
                 26 
                 9 
                 8.78 
                 5.78 
               
               
                 Cluster 2 
               
               
                 Methyl 
                 0 
                 16 
                 2 
                 2.49 
                 2.91 
                 0 
                 25 
                 5 
                 5.75 
                 5.53 
               
               
                 Cluster 3 
               
               
                   
               
            
           
         
       
     
     Properties of 6 mA distribution in nucleosome linkers. In  Oxytricha , methyl cluster 1=between 5′ chromosome end and +1 nucleosome; methyl cluster 2=between +1 and +2 nucleosome; methyl cluster 3=between +2 and +3 nucleosome. In  Tetrahymena , methyl cluster 1=between +1 and +2 nucleosome; methyl cluster 2=between +2 and +3 nucleosome; methyl cluster 3=between +3 and +4 nucleosome. Consensus +1/+2/+3/+4 nucleosome positions: 193, 402, 618, 837 bp downstream of  Oxytricha  5′ chromosome ends; 112, 304, 497, 698 bp downstream of  Tetrahymena  TSSs. 
     Example 3 
     Purification and Reconstitution of the Ciliate 6 mA Methyltransferase, MTA1c 
     To uncover the functions of 6 mA in vivo, we set out to identify and disrupt putative 6 mA methytransferases (MTases). The  Oxytricha  genome encodes a large number of candidate methyltransferases (Table 5), rendering it impractical to test gene function, one at a time or in combination. To identify the ciliate 6 mA MTase, we undertook a biochemical approach by fractionating nuclear extracts and identifying candidate proteins that co-purified with DNA methylase activity. The organism of choice for this experiment was  Tetrahymena thermophila , a ciliate that divides significantly faster than  Oxytricha  (˜2 h versus 18 h; Cassidy-Hanley, 2012; Laughlin et al., 1983). This faster growth time rendered it feasible to culture large amounts of  Tetrahymena  cells for nuclear extract preparation.  Tetrahymena  and  Oxytricha  exhibit similar genomic localization and 6 mA abundance ( FIGS. 8A-8B and 9A-9F ). We thus reasoned that the enzymatic machinery responsible for 6 mA deposition is conserved between  Tetrahymena  and  Oxytricha , and that  Tetrahymena  could serve as a tractable biochemical system for identifying the ciliate 6 mA MTase. 
     We prepared nuclear extracts from log-phase  Tetrahymena  cells, since 6 mA could be readily detected at this developmental stage through quantitative MS and PacBio sequencing ( FIGS. 8A-8B and 9A-9F ). Nuclear extracts were incubated with radiolabeled S-adenosyl-L-methionine (SAM) and PCR-amplified DNA substrate to assay for DNA methylase activity. Passage of the nuclear extract through an anion exchange column resulted in the elution of two distinct peaks of DNA methylase activity, both of which were heat sensitive ( FIGS. 2C, 10A, and 10B ). Western blot analysis confirmed that both peaks of activity mediate methylation on 6 mA ( FIG. 10C ). The resulting fractions were further purified and subjected to MS. Only four proteins-termed MTA1, MTA9, p1, and p2-were detected at higher abundance in fractions with high DNA methylase activity ( FIGS. 2C and 2D ). p1 and p2 contain homeobox-like domains, suggesting a DNA binding function for an undetermined process ( FIG. 10D ). On the other hand, MTA1 and MTA9 are both MT-A70 proteins. Such domains are widely known to mediate m6A RNA methylation in eukaryotes (Liu et al., 2014). MTA1 and MTA9 received the large majority of peptide matches, relative to all other MT-A70 genes encoded by the  Tetrahymena  genome ( FIG. 2D ; Table 6). Curiously, although poly(A)-selected RNA transcripts were present from all MT-A70 genes ( FIG. 2D ), almost all peptides in fractions with high DNA methylase activity corresponded to MTA1 and MTA9. The poly(A) +  RNA expression profiles of MTA1, MTA9, p1, and p2 are remarkably similar ( FIG. 9K ), peaking early in the sexual cycle. This coincides with a sharp increase in nuclear 6 mA, as evidenced from immunostaining (Wang et al., 2017). Accumulation of MTA1, MTA9, p1, and p2 therefore correlates with the presence of 6 mA in vivo. 
     We next investigated the phylogenetic relationship of MTA1 and MTA9 to other eukaryotic MT-A70 domain-containing proteins. Two widely studied mammalian MT-A70 proteins—METTL3 and METTL14 (Ime4 and Kar4 in yeast)-form a heterodimeric complex that is responsible for m6A methylation on mRNA. METTL3 is the catalytically active subunit, while METTL14 functions as an RNA-binding scaffold protein (Sledi arid Jinek, 2016; Wang et al., 2016a, 2016b). MTA1 and MTA9 derive from distinct monophyletic clades, outside of those that contain mammalian METTL3, METTL14, and  C. elegans  DAMT-1 (METTL4) ( FIG. 2A ). Thus, MTA1 and MTA9 are divergent MT-A70 family members that are phylogenetically distinct from all previously studied RNA and DNA N6-methyladenine MTases. We then asked whether MTA1 and MTA9 are also present in other eukaryotes with a similar occurrence of 6 mA in ApT motifs as  Tetrahymena . We queried the genomes of  Oxytricha , green algae, and eight basal yeast species, all of which exhibit this distinct methylation pattern (as evidenced from  FIGS. 1A-1E ;  FIGS. 9A-9E ; Fu et al., 2015; Mondo et al., 2017). For all of these taxa, we can identify MT-A70 homologs that are monophyletic with MTA1 and MTA9 ( FIG. 2B ). On the other hand, MT-A70 homologs from multicellular eukaryotes, including  Arabidopsis, C. elegans, Drosophila , and mammals, grouped exclusively with METTL3, METTL14, and METTL4 lineages, but not MTA1 or MTA9. None of these latter genomes exhibit a consensus ApT dinucleotide methylation motif for 6 mA (Greer et al., 2015; Koziol et al., 2016; Liang et al., 2018; Liu et al., 2016; Wu et al., 2016; Xiao et al., 2018; Zhang et al., 2015). We note that the absence of an ApT dinucleotide motif is based on data from a limited number of cell types, developmental stages, and culture conditions tested in these studies. Nonetheless, within the scope of currently available data, the presence of MTA1 and MTA9 correlates with the distinctive genomic localization of 6 mA within ApT motifs. 
     We then sought to determine whether MTA1 and/or MTA9 are bona fide 6 mA methyltransferases. MTA1, but not MTA9, contains a catalytic DPPW motif ( FIG. 10E )—a hallmark of N6-adenosine methyltransferases (Iyer et al., 2016). Surprisingly, recombinant full-length  Tetrahymena  MTA1 and MTA9 ( FIG. 10G ) showed no detectable DNA methyltransferase activity, individually or together ( FIG. 2E ). Examination of the MTA1 and MTA9 sequences revealed that neither protein possesses a predicted nucleic acid binding domain ( FIG. 10D ). In contrast, METTL3, which catalyzes m6A methylation on RNA, contains two tandem CCCH-type zinc finger motifs, necessary for RNA binding (Huang et al., 2019; Wang et al., 2016a). Additional co-factors may thus be necessary for MTA1/7 to engage DNA substrates. Indeed, the p1 and p2 proteins that co-elute with MTA1/7 in nuclear extracts possess homeobox-like domains predicted to bind DNA. We then tested whether these accessory factors, in addition to MTA1/7, are necessary for 6 mA methylation. Strikingly, mixing recombinant, full-length p1, p2, MTA1, and MTA9 resulted in robust 6 mA methylation in vitro ( FIGS. 2E and 2F ). This activity was abolished when each protein was omitted, indicated that all four are necessary for 6 mA methylation. Furthermore, MTA1 harboring a D209A mutation in the catalytic DPPW motif showed no activity, even in the presence of MTA9, p1, and p2 ( FIG. 2E ). We also created double mutations in MTA1 (N370A, E371A), which lie in the conserved region that interacts with the 2′ and 3′-hydroxyl groups of the ribose moiety in the SAM cofactor ( FIG. 2E ). This mutant protein also exhibited no 6 mA methylase activity. Taken together, we find that four proteins—MTA1, MTA9, p1, and p2—are necessary for 6 mA methylation in vitro, with MTA1 the likely catalytic subunit. Henceforth, we refer to these four proteins as the putative MTA1 complex (MTA1c). 
     Purification of the MTA1c proteins from an  E. coli  overexpression system raises the possibility of methyltransferase activity arising from contaminating Dam methylase; however, we exclude this possibility for three reasons. (1) The DNA substrate used in this assay does not contain 5′-NATC-3′ sites, which are recognized and methylated by Dam methylase (Horton et al., 2006). (2) Methyltransferase activity was only observed when all four recombinant proteins were incubated with DNA. If contaminating Dam methylase were present in one or more of these protein preparations, then background activity should be observed when subsets of these proteins are used in the assay. 3) Mutation of MTA1 catalytic residues leads to loss of methylation, which is also inconsistent with contaminating methyltransferase activity. 
                     TABLE 5               Candidate genes in ciliates.                  MT-A70 genes in Oxytricha trifallax                                     Gene name in               UniProt ID   this study   OxyDB gene name                       J9IF92_9SPIT   MTA1   Contig12701.0.0.g16           J9IGS7_9SPIT   TAMT-1   Contig17486.0.g100           J9J9V7_9SPIT   MTA1-B   Contig16314.0.g25           J9HW68_9SPIT   MTA9   Contig1237.1.g126           J9IMU5_9SPIT   MTA9-B   Contig17413.0.g36                             MT-A70 genes in Tetrahymena thermophila                             Gene name in   Tetrahymena Genome       UniProt ID   this study   Database gene name               Q22GC0_TETTS   MTA1   TTHERM_00704040       Q23TW8_TETTS   MTA2   TTHERM_00962190       I7LVP8_TETTS   MTA3/TAMT-1-B   TTHERM_00136470       I7MGX6_TETTS   MTA4   TTHERM_00558100       Q23RE0_TETTS   MTA5/TAMT-1   TTHERM_00388490       I7MIF9_TETTS   MTA9   TTHERM_00301770       Q22XT1_TETTS   MTA9-B   TTHERM_01005150                         METTL16 homologs in Oxytricha trifallax                             UniProt ID   OxyDB gene name                       J9F3J7_9SPIT   Contig11945.0.g48           J9J5P9_9SPIT   Contig7462.0.g41           J9III0_9SPIT   Contig4244.0.g39                             N6AMT1 homologs in Oxytricha trifallax                             UniProt ID   OyDB gene name                       J9IFV1_9SPIT   Contig7751.0.g12                             Accessory factor genes in Tetrahymena thermophila                                     Gene name in   Tetrahymena Genome           UniProt ID   this study   Database gene name                       Q22VV9_TETTS   p1   TTHERM_00161750           I7M8B9_TETTS   p2   TTHERM_00439330                             ISWI homologs in Oxytricha trifallax and Tetrahymena thermophile                                 Tetrahymena Genome       UniProt ID   OxyDB gene name   Database gene name               I7M280_TETTS       TTHERM_00137610       J9FBJ2_9SPIT   Contig11737.0.g12                    
The Uniprot ID of each gene is listed. The  Oxytricha  macronuclear genome encodes five genes belonging to the MT-A70 family (Iyer et al., 2016; Swart et al., 2013). Such genes commonly function as RNA m6 A MTases in eukaryotes, having evolved from m.MunI-like MTases in bacterial restriction-modification systems (Iyer et al., 2016). An MT-A70 gene belonging to the METTL4 subclade, DAMT1, is a putative 6 mA methyltransferase in  C. elegans  (Greer et al., 2015). However, none of the  Oxytricha  MT-A70 genes in this Table cluster together with METTL4 on a phylogenetic tree ( FIGS. 2A and 9G ). The  Oxytricha  genome also contains homologs of a structurally distinct RNA m6 A MTase, METTL16, which was reported to methylate U6 snRNA (Table 5) (Pendleton et al., 2017; Warda et al., 2017). Another candidate, N6AMT1—which does not contain an MT-A70 domain—was recently found to mediate DNA 6 mA methylation in human cells (Xiao et al., 2018). An N6AMT1 homolog is also present in the  Oxytricha  genome. Accessory factors refer to the p1 and p2 proteins, which are necessary for 6 mA methylation by MTA1 and MTA9 in vitro. The UniProt IDs of putative ISWI homologs in  Oxytricha  and  Tetrahymena  are also listed.
 
                     TABLE 6               Mass spectrometry analysis of MTA1, MTA9, p1, and p2 proteins.                  Data from Low Salt Fraction                             Gene name in   % of protein covered by peptide       UniProt ID   this study   data from LC-MS/MS experiment               Q22GC0_TETTS   MTA1   78.8%       I7MIF9_TETTS   MTA9   46.3%       Q22W9_TETTS   p1   41.9%       I7M8B9_TETTS   p2   81.7%                         Data from High Salt Fraction                             Gene name in   % of protein covered by peptide       UniProt ID   this study   data from LC-MS/MS experiment               Q22GC0_TETTS   MTA1   69.9%       I7MIF9_TETTS   MTA9   72.2%       Q22VV9_TETTS   p1   55.3%       I7M8B9_TETTS   p2   93.4%                    
Percentage of each polypeptide that is covered by peptide data is calculated. “Low Salt Sample” and “High Salt Sample” correspond to partially purified nuclear extracts that elute as two distinct peaks of activity from a Q sepharose anion exchange column ( FIG. 2C ).
 
     Example 4 
     MTA1c Preferentially Methylates ApT Dinucleotides in dsDNA 
     We next investigated the substrate preferences of MTA1c. First, in vitro transcription was performed to generate doublestranded RNA (dsRNA) and single-stranded RNA (ssRNA) from the input dsDNA substrate. We found that MTA1c methylates dsDNA but not dsRNA or ssRNA of the same sequence, indicating that it is selective for DNA over RNA ( FIG. 10H ). We then generated a series of dsDNA substrates by annealing oligonucleotide pairs of different length and sequence. All of these substrates are bona fide  Tetrahymena  genomic DNA sequences. In each case, MTA1c can methylate the annealed dsDNA but not ssDNA ( FIGS. 2G and 10I ). 
     Since 6 mA methylation mainly lies in ApT dinucleotides in vivo ( FIGS. 1D and 9D ), we asked whether MTA1c preferentially methylates this motif. To test this, we used a 27 bp dsDNA substrate with two ApT dinucleotides in its native sequence ( FIG. 2G ). We disrupted one or both ApT motifs ( FIG. 2G ) by mutually swapping the 5′ A with a neighboring base 5′-CAT-3′→5′-ACT-3′. Disrupting both ApT dinucleotides resulted in &gt;10-fold reduced methylation, while disrupting only one motif led to a 2- to 4-fold loss ( FIGS. 2G and 10K ). 
     Given that 6 mA occurs on both strands of genomic DNA in vivo ( FIGS. 1E and 9E ), we asked whether pre-existing methylation of one strand affects MTA1c activity. DNA oligonucleotides were nonspecifically methylated with 6 mA using EcoGII (Murray et al., 2018), a bacterial 6 mA methyltransferase. After rigorous purification, samples were annealed to an unmethylated, complementary strand to yield hemimethylated dsDNA ( FIG. 10F ). MTA1c activity was 3- to 3.5-fold higher on hemimethylated substrates, relative to unmethylated dsDNA ( FIG. 2G ). This effect was similar between dsDNA substrates pre-methylated on the sense or antisense strand, consistent with the lack of an overt strand bias in 6 mA locations in vivo ( FIGS. 1E and 9E ). Importantly, the increase in MTA1c activity cannot be attributed to contaminating EcoGII in hemimethylated substrates, since no activity was observed in the absence of MTA1c ( FIG. 10J ). Thus, pre-existing 6 mA methylation stimulates MTA1c, indicative of a positive feedback loop. 
     We then asked whether MTA1c activity is modulated not only by the dinucleotide motif sequence per se, but also by flanking sequences. This may manifest as the wide variation in frequency of DNA 4-mer containing a methylated ApT dinucleotide 5′-NA*TN-3′ in vivo ( FIG. 10M ). To test this, we used a dsDNA substrate containing two ApT dinucleotides, both within a 5′-CATT-3′. Swapping of the ApT motif with the adjacent downstream DNA residue produced substrates containing 5′-TATA-3′ ( FIG. 10L ). Substrates with this change at both locations had 4-fold less MTA1c activity, and an intermediate effect when only one dinucleotide was altered ( FIG. 10L ). These data indicate that 5′-CATT-3′ is the preferred methylation substrate, consistent with the higher frequency of methylated 5-CA*TT-3′ versus 5-TA*TA-3′ in both  Tetrahymena  and  Oxytricha  genomic DNA ( FIG. 10M ). The difference in frequency of methylated sequences cannot simply be attributed to the higher frequency of the 4nt 5′-CATT-3′ motif versus 5′-TATA-3′ in the genome, because the opposite trend is observed ( FIG. 10N ). Thus, MTA1c is sensitive to variation in DNA sequences flanking the ApT dinucleotide motif. 
     Example 5 
     MTA1 is Necessary for 6 mA Methylation In Vivo 
     Having established that MTA1c is a 6 mA methyltransferase, we tested the role of MTA1c in mediating 6 mA methylation in vivo in  Oxytricha , for which we have ease of generating mutants. The genome-wide localization of 6 mA is conserved between  Oxytricha  and  Tetrahymena  ( FIGS. 1A-1E and 9A-9F ), implying similar underlying enzymatic machinery. Indeed, all four component genes—MTA1, MTA9, p1, and p2—are clearly conserved between both species ( FIGS. 9G-9J ). The DPPW catalytic motif is also completely conserved in  Tetrahymena  and  Oxytricha  MTA1 but not MTA9, suggesting that MTA1 is the likely catalytic subunit of MTA1c in both ciliates ( FIG. 10E ). To abrogate MTA1c function, we disrupted the  Oxytricha  MTA1 gene by inserting an ectopic DNA sequence 49 bp downstream of the start codon, resulting in a frameshift mutation and loss of the C-terminal MTase domain ( FIG. 3A ).  Oxytricha  has two MTA1 paralogs, named MTA1 and MTA1-B ( FIGS. 2A and 9G ). We focused on MTA1 because MTA1-B is not expressed in vegetative  Oxytricha  cells (Swart et al., 2013), which we used to profile 6 mA locations via SMRT-seq. Dot blot analysis confirmed a significant reduction in bulk 6 mA levels in mutant lines ( FIG. 3B ). We then examined 6 mA positions at high resolution using SMRT-seq to understand how the DNA methylation landscape is altered in mta1 mutants. Notably, these mutants exhibit genome-wide loss of 6 mA, with complete abolishment of the dimethylated ApT motif, and reduction in frequency of all other methylated dinucleotide motifs ( FIGS. 3C-3E ). These findings are consistent across all biological replicates and are robust to wide variation in SMRT-seq parameters for calling 6 mA modifications ( FIGS. 11B-11D ). It cannot be attributed to variation in sequencing coverage between wild-type and mutant lines. The loss of methylated ApT dinucleotides in mta1 mutants is consistent with our in vitro data suggesting that MTA1c primarily methylates ApT sites ( FIGS. 2G and 10K ). The Inter Pulse Duration ratio (degree of polymerase slowing during PacBio sequencing due to presence of a modified base) and estimated fractional methylation also decreased significantly at called 6 mA sites in mta1 mutants (p&lt;2.2×10-16, Wilcoxon rank-sum test) ( FIG. 11A ). MTA1 is therefore necessary for a significant proportion of in vivo 6 mA methylation events in  Oxytricha.    
     What are the phenotypic consequences of 6 mA loss in vivo? It has been proposed that DNA methylation—including 6 mA and cytosine methylation—is involved in nucleosome organization (Fu et al., 2015; Huff and Zilberman, 2014). We thus asked whether nucleosome organization is altered in mta1 mutants. We quantified nucleosome “fuzziness,” defined as the SD of MNase-seq read locations surrounding the called nucleosome peak (Lai and Pugh, 2017; Mavrich et al., 2008). A poorly positioned nucleosome consists of a shallow and wide peak of MNase-seq reads, manifested by a high fuzziness score. Nucleosomes were first grouped according to the change in flanking 6 mA between wild-type and mta1 mutant cells ( FIGS. 12A-12G ). The nucleosomes that experience large changes in flanking 6 mA exhibit significantly greater increase in fuzziness, compared to nucleosomes with little change in flanking 6 mA ( FIGS. 12A and 12D ). Such nucleosomes also exhibit changes in occupancy that are consistent with an increase in fuzziness ( FIGS. 12A and 12E ). These results are robust to variation in MNase digestion ( FIGS. 14C and 14D ). On the other hand, nucleosome linkers do not change in length or occupancy, even though 6 mA is lost from these regions ( FIGS. 12B, 12C, 12F, and 12G ). We conclude that 6 mA exerts subtle effects on nucleosome organization in vivo. 
     Example 6 
     6 mA Disfavors Nucleosome Occupancy Across the Genome In Vitro but not in Vivo 
     Multiple factors, including 6 mA, DNA sequence, and chromatin remodeling complexes, may collectively contribute to nucleosome organization in vivo. The effect of 6 mA could therefore be masked by these elements. We next sought to determine whether 6 mA directly impacts nucleosome organization. To this end, we assembled chromatin in vitro using  Oxytricha  gDNA, which contains cognate 6 mA. To obtain a matched negative control lacking DNA methylation, 98 complete chromosomes were amplified using PCR ( FIG. 4A ), purified and subsequently mixed together in stoichiometric ratios to obtain a “mini-genome” ( FIG. 4B ). These chromosomes collectively reflect overall genome properties, including AT content, chromosome length, and transcriptional activity (Table 7). Native genomic DNA (containing 6 mA) and amplified mini-genome DNA (lacking 6 mA) were each assembled into chromatin in vitro using  Xenopus  or  Oxytricha  histone octamers ( FIGS. 13A-13F ) and analyzed using MNase-seq. We computed nucleosome occupancy from the native genome and mini-genome samples across 199,795 overlapping DNA windows, spanning all base pairs in the 98 chromosomes. This allowed the direct comparison of nucleosome occupancy in each window of identical DNA sequence, with and without 6 mA ( FIGS. 4C and 4D ). Windows exhibit lower nucleosome occupancy with increasing 6 mA, confirming the quantitative nature of this effect. Furthermore, similar trends were observed for both native  Oxytricha  and recombinant  Xenopus  histones, suggesting that the effects of 6 mA on nucleosome organization arise mainly from intrinsic features of the histone octamer rather than from species-specific variants ( FIGS. 4C and 4D ). These results are also robust to the extent of MNase digestion of reconstituted chromatin ( FIG. 14A ). 
     We then directly compared the impact of 6 mA on nucleosome occupancy in vitro and in vivo. Loss of 6 mA in vitro is achieved by mini-genome construction, while loss in vivo is achieved by the mta1 mutation. For each overlapping DNA window, we calculated the difference in nucleosome occupancy: (1) between native genome and mini-genome DNA in vitro, and (2) between wild-type and mta1 mutants in vivo ( FIG. 4C ). Nucleosome occupancy is indeed lower in the presence of 6 mA methylation in vitro ( FIGS. 4C and 4D ). In contrast, no change in nucleosome occupancy is observed in vivo ( FIGS. 4C and 4E ). This result is consistent with our earlier analysis of linker occupancy in mta1 mutants ( FIGS. 12C and 12G ). We note that highly methylated DNA windows show greater change in 6 mA relative to mta1 mutants ( FIG. 3D ). Yet, these windows do not change in nucleosome occupancy in vivo. We conclude that 6 mA methylation locally disfavors nucleosome occupancy in vitro, but that this intrinsic effect can be overcome by endogenous chromatin factors in vivo. 
                     TABLE 7                  Descriptive statistics of reference genomes.                             Native genomic DNA   Mini-genome DNA                                             Chromosome   2449 +/− 742   2107 +/− 778            length (bp)   Min = 1155   Min = 1201               Max = 6494   Max = 4659           SMRT-seq    177.4 +/− 117.0   205.3 +/− 136.1           coverage (x)   Min = 75.1   Min = 77.8               Max = 1392.6   Max = 918.4           Total number   46,322   2,344           of 6mA marks           in genome           6mA sites per   12 +/− 8   24 +/− 16           chromosome   Min = 0   Min = 0               Max = 73   Max = 73           AT content (%)   67.8 +/− 3.0   66.5 +/− 2.7                Min = 55.7   Min = 60.2               Max = 76.2   Max = 72.2           RNAseq    34.4 +/− 75.2   53.7 +/− 71.5           (FPKM)   Min = 0.0   Min = 0.1               Max = 1444.5   Max = 424.8                        
Properties of  Oxytricha  chromosomes in native genomic DNA and mini-genome DNA. “+/−” indicates one standard deviation above or below the mean.
 
     Example 7 
     Modular Synthesis of Epigenetically Defined Chromosomes 
     The above experiments used kinetic signatures from SMRT-seq data to infer the presence of 6 mA marks in genomic DNA. We next sought to confirm that 6 mA is directly responsible for disfavoring nucleosomes in vitro, and to understand how this effect could be overcome by cellular factors. 6 mA-containing oligonucleotides were annealed and subsequently ligated with DNA building blocks to form full-length chromosomes. Importantly, these chromosomes contain 6 mA at all locations identified by SMRT-seq in vivo. The representative chromosome, Contig1781.0, is 1.3 kb, contains a clearly defined TSS, and encodes a single highly transcribed gene with a predicted RING finger domain. The length and gene structure are characteristic of typical  Oxytricha  chromosomes ( FIG. 5A ). We independently validated the location of 6 mA in vivo by sequencing chromosomal DNA immunoprecipitated with an anti-6 mA antibody ( FIG. 5A ). 
     Four chromosome variants were synthesized, with cognate 6 mA sites on neither, one, or both DNA strands (chromosomes 1-4 in  FIGS. 5B and 5C ). Chromatin was assembled by salt dialysis with either  Oxytricha  or  Xenopus  nucleosomes and subsequently digested with MNase to obtain mononucleosomal DNA ( FIGS. 6A and 13G ). Tiling qPCR was used to quantify nucleosome occupancy at ˜50 bp increments along the entire length of the synthetic chromosome ( FIG. 6B ). The fully methylated locus exhibits a ˜46% reduction in nucleosome occupancy relative to the unmethylated variant, while hemimethylated chromosomes containing half the number of 6 mA marks showed intermediate nucleosome occupancy at the corresponding region ( FIG. 6B ). The reduction in nucleosome occupancy was confined to the methylated region and not observed across the rest of the chromosome. Similar trends were observed when chromatin was assembled using the NAP1 histone chaperone ( FIG. 14F . top panel). indicating that this effect is not an artifact of the salt dialysis method. Furthermore, moving 6 mA to an ectopic location (chromosome 5 in  FIGS. 5B and 5C ) decreases nucleosome occupancy at that site ( FIG. 6C ). We conclude that 6 mA directly disfavors nucleosome occupancy in a local, quantitative manner in vitro. 
     Example 8 
     Chromatin Remodelers Restore Nucleosome Occupancy Over 6 mA Sites 
     Nucleosome occupancy in vivo is influenced not only by DNA sequences but also by trans-acting factors such as ATP-dependent chromatin remodeling factors (Struhl and Segal, 2013). We used synthetic, methylated chromosomes to test how the well-studied chromatin remodeler ACF responds to 6 mA in native DNA. ACF generates regularly spaced nucleosome arrays in vitro and in vivo (Clapier and Cairns, 2009; Ito et al., 1997). Its catalytic subunit ISWI is conserved across eukaryotes, including  Oxytricha  and  Tetrahymena  (Table 5). Synthetic chromosomes were assembled into chromatin by salt dialysis as before and then incubated with ACF in the presence of ATP ( FIGS. 13H and 6D ). We find that ACF partially—but not completely—restores nucleosome occupancy over the methylated locus in an ATP-dependent manner ( FIG. 6D ). This effect is observed when ACF was added to chromatin assembled by salt dialysis or the NAP1 histone chaperone ( FIGS. 6D and 14F ). ACF also restores nucleosome occupancy over methylated loci in native genomic DNA ( FIGS. 6E and 13I ), indicating that the effect is not restricted to a single chromosome. This result is robust to the extent of MNase digestion ( FIG. 14B ). Although the heterologous system used here may differ from endogenous chromatin assembly factors in  Oxytricha , our experiment illustrates the principle that trans-acting factors can counteract or even overcome the effect of 6 mA on nucleosome organization. 
     Example 9 
     Disruption of MTA1 Impacts Gene Expression and Sexual Development 
     Since mta1 mutants exhibit genome-wide loss of 6 mA, we assayed these cells for transcriptional changes by poly(A) +  RNAseq. Only a small minority of genes show significant changes in gene expression (10% false discovery rate [FDR];  FIG. 7A ). To examine the methylation status of these differentially expressed genes, we grouped them according to “starting” methylation level, as defined by the total number of 6 mA marks near the TSS in wild-type cells. Genes exhibit two distinct transcriptional responses: those with low starting levels of 6 mA exhibit a small change in 6 mA between wild-type and mutant cells ( FIG. 3D ) and tend to be significantly upregulated in mutant lines (p=2.8×10 −9 , Fisher&#39;s exact test;  FIG. 7B ). Surprisingly, genes with high starting 6 mA are not enriched in differentially expressed genes (p&gt;0.1, Fisher&#39;s exact test), even though they exhibit greater loss of 6 mA in mutants ( FIG. 3D ). Steady-state RNA-seq levels are therefore largely robust to drastic changes in 6 mA levels. Since most, but not all, 6 mA is lost from mta1 mutants ( FIG. 3C ), it is also possible that residual DNA methylation across the genome sufficiently buffers genes from changes in transcription. 
     Because the aforementioned phenotypic changes were assayed in vegetative  Oxytricha  cells, we asked whether MTA1 may play roles outside of this developmental state. MTA1 transcript levels are markedly upregulated in the sexual cycle, as assayed by poly(A). RNA-seq ( FIG. 7C ). Strikingly, mta1 mutants fail to complete the sexual cycle when induced to mate and display complete lethality ( FIG. 7D ). Our data do not exclude the possibility that m6A RNA methylation, in addition to 6 mA DNA methylation, is also impacted by MTA1 loss during development. Further studies would clarify the role of MTA1 in these pathways. 
     Example 10 
     Discussion 
     The present disclosure has identified MTA1c as a conserved, hitherto undescribed 6 mA methyltransferase. It consists of two MT-A70 proteins (MTA1/MTA9) and two homeobox-like proteins (p1/p2). The composition of MTA1c provides immediate insights into how it specifically methylates DNA ( FIG. 7F ). MTA1 likely mediates transfer of the methyl group from SAM to the acceptor adenine moiety, given that it contains conserved amino acid residues implicated in catalysis and SAM binding ( FIG. 10E ). Indeed, we show that these residues are necessary for its activity ( FIG. 2E ). While MTA1 constitutes the catalytic center, it lacks a CCCH-type zinc finger domain that is necessary for RNA binding in the canonical m6A methyltransferase METTL3. Instead, nucleic acid binding is likely assumed by the homeobox-like domains in p1 and p2, which are known to specifically engage dsDNA through helix-turn-helix motifs. 
     The observation that MTA1c is more active in the presence of pre-methylated DNA templates is reminiscent of the CpG methyltransferase DNMT1. Yet, MTA1c and DNMT1 exhibit distinct protein domain architectures. Further biochemical studies are required to elucidate the molecular basis of this property. A distinct MT-A70 protein, named TAMT-1, was recently reported to act as a 6 mA methyltransferase in  Tetrahymena , (Luo et al., 2018), suggesting that multiple enzymes mediate 6 mA deposition. It remains to be determined how MTA1c and TAMT-1 collectively mediate DNA methylation at various developmental stages, and whether cross-talk occurs between these enzymes. 
     In addition to identifying the ciliate 6 mA methyltransferase, we investigated the function of 6 mA in vitro by building epigenetically defined chromosomes. We show that 6 mA directly disfavors nucleosome occupancy in a local, quantitative manner, independent of DNA sequence ( FIG. 7E ). Our experiments do not reveal exactly how 6 mA disfavors nucleosome occupancy. Early studies suggest that 6 mA destabilizes dA:dT base pairing, leading to a decrease in the melting temperature of DNA (Engel and von Hippel, 1978). Whether this or some other property of 6 mA contributes to lowered nucleosome stability awaits further investigation. 
     Intriguingly, nucleosome organization exhibits only subtle changes after genome-wide loss of 6 mA ( FIG. 7E ). Only a small set of genes (&lt;10%) is transcriptionally dysregulated. It is possible that residual 6 mA in mta1 mutants could mask relevant phenotypes. Nonetheless, our results caution against interpreting 6 mA function solely based on correlation with genomic elements. We also find that 6 mA intrinsically disfavors nucleosomes in vitro, but—crucially—this effect can be overridden by distinct factors in vitro and in vivo. We propose that phased nucleosome arrays are first established in vivo, which then restrict MTA1-mediated methylation to linker regions due to steric hindrance. This in turn decreases the fuzziness of flanking nucleosomes, reinforcing chromatin organization. Therefore, 6 mA tunes nucleosome organization in vivo. Our data do not support the hypothesis that nucleosome phasing is established by predeposited 6 mA. 
     More broadly, our work showcases the utility of  Oxytricha  chromosomes for advancing chromatin biology. By extending current technologies (Muller et al., 2016), it should be feasible to introduce both modified nucleosomes and DNA methylation in a site-specific manner on full-length chromosomes. Such “designer” chromosomes will serve as powerful tools for studying DNA-templated processes such as transcription within a fully native DNA environment. 
     REFERENCES 
     The following references, to the extent that they provide exemplary procedural or other details supplementary to those set forth herein, are specifically incorporated herein by reference.
     Altschul, S. F., Madden, T. L., Schaffer, A. A., Zhang, J., Zhang, Z., Miller, W., and Lipman, D. J. (1997). Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25, 3389-3402.   Ammermann, D., Steinbruck, G., Baur, R., and Wohlert, H. (1981). Methylated bases in the DNA of the ciliate  Stylonychia mytilus . Eur. J. Cell Biol. 24, 154-156.   An, W., and Roeder, R. G. (2004). Reconstitution and transcriptional analysis of chromatin in vitro. Methods Enzymol. 377, 460-474.   Batut, P., Dobin, A., Plessy, C., Carninci, P., and Gingeras, T. R. (2013). Highfidelity promoter profiling reveals widespread alternative promoter usage and transposon-driven developmental gene expression. Genome Res. 23, 169-180.   Beh, L. Y., Muller, M. M., Muir, T. W., Kaplan, N., and Landweber, L. F. (2015). DNA-guided establishment of nucleosome patterns within coding regions of a eukaryotic genome. Genome Res. 25, 1727-1738.   Beh et al., Identification of a DNA N6-Adenine Methyltransferase Complex and Its Impact on Chromatin Organization, Cell (2019), https://doi.org/10.1016/j.ce11.2019.04.028.   Bern, M., Kil, Y. J., and Becker, C. (2012). Byonic: Advanced Peptide and Protein Identification Software. Curr. Protoc. Bioinformatics. 13, 13.20.   Blankenberg, D., Von Kuster, G., Coraor, N., Ananda, G., Lazarus, R., Mangan, M., Nekrutenko, A., and Taylor, J. (2010). Galaxy: a web-based genome analysis tool for experimentalists. Curr. Protoc. Mol. Biol. 19, 19.10.1-19.10.21.   Bracht, J. R., Fang, W., Goldman, A. D., Dolzhenko, E., Stein, E. M., and Landweber, L. F. (2013). Genomes on the edge: programmed genome instability in ciliates. Cell 152, 406-416.   Bromberg, S., Pratt, K., and Hattman, S. (1982). Sequence specificity of DNA adenine methylase in the protozoan  Tetrahymena thermophila . J. Bacteriol. 150, 993-996.   Brownell, J. E., Zhou, J., Ranalli, T., Kobayashi, R., Edmondson, D. G., Roth, S. Y., and Allis, C. D. (1996).  Tetrahymena  histone acetyltransferase A: a homolog to yeast Gcn5p linking histone acetylation to gene activation. Cell 84, 843-851.   Cassidy-Hanley, D. M. (2012).  Tetrahymena  in the Laboratory: Strain Resources, Methods for Culture, Maintenance, and Storage. Methods Cell Biol. 109, 237-276.   Chen, X., Bracht, J. R., Goldman, A. D., Dolzhenko, E., Clay, D. M., Swart, E. C., Perlman, D. H., Doak, T. G., Stuart, A., Amemiya, C. T., et al. (2014). The architecture of a scrambled genome reveals massive levels of genomic rearrangement during development. Cell 158, 1187-1198.   Clapier, C. R., and Cairns, B. R. (2009). The biology of chromatin remodeling complexes. Annu. Rev. Biochem. 78, 273-304.   Cummings, D. J., Tait, A., and Goddard, J. M. (1974). Methylated bases in DNA from Paramecium aurelia. Biochim. Biophys. Acta 374, 1-11.   Debelouchina, G. T., Gerecht, K., and Muir, T. W. (2017). Ubiquitin utilizes an acidic surface patch to alter chromatin structure. Nat. Chem. Biol. 13, 105-110.   Eisen, J. A., Coyne, R. S., Wu, M., Wu, D., Thiagarajan, M., Wortman, J. R., Badger, J. H., Ren, Q., Amedeo, P., Jones, K. M., et al. (2006). Macronuclear genome sequence of the ciliate  Tetrahymena thermophila , a model eukaryote. PLoS Biol. 4, e286.   Eng, J. K., McCormack, A. L., and Yates, J. R. (1994). An approach to correlate tandem mass spectral data of peptides with amino acid sequences in a protein database. J. Am. Soc. Mass Spectrom. 5, 976-989.   Engel, J. D., and von Hippel, P. H. (1978). Effects of methylation on the stability of nucleic acid conformations. Studies at the polymer level. J. Biol. Chem. 253, 927-934.   Fang, W., Wang, X., Bracht, J. R., Nowacki, M., and Landweber, L. F. (2012). Piwi-interacting RNAs protect DNA against loss during  Oxytricha  genome rearrangement. Cell 151, 1243-1255.   Finn, R. D., Clements, J., Arndt, W., Miller, B. L., Wheeler, T. J., Schreiber, F., Bateman, A., and Eddy, S. R. (2015). HMMER web server 2015 update. Nucleic Acids Res. 43 (W1), W30-W38.   Fioravanti, A., Fumeaux, C., Mohapatra, S. S., Bompard, C., Brilli, M., Frandi, A., Castric, V., Villeret, V., Viollier, P. H. P., and Biondi, E. G. (2013). DNA binding of the cell cycle transcriptional regulator GcrA depends on N6-adenosine methylation in  Caulobacter crescentus  and other Alphaproteobacteria. PloS Genet. 9, e1003541.   Fu, Y., Luo, G.-Z., Chen, K., Deng, X., Yu, M., Han, D., Hao, Z., Liu, J., Lu, X., Dore, L. C., et al. (2015). N6-methyldeoxyadenosine marks active transcription start sites in  Chlamydomonas . Cell 161, 879-892.   Fyodorov, D. V., and Kadonaga, J. T. (2003). Chromatin assembly in vitro with purified recombinant ACF and NAP-1. Methods Enzymol. 371, 499-515.   Giardine, B., Riemer, C., Hardison, R. C., Burhans, R., Elnitski, L., Shah, P., Zhang, Y., Blankenberg, D., Albert, I., Taylor, J., et al. (2005). Galaxy: a platform for interactive large-scale genome analysis. Genome Res. 15, 1451-1455.   Goecks, J., Nekrutenko, A., and Taylor, J.; Galaxy Team (2010). Galaxy: a comprehensive approach for supporting accessible, reproducible, and transparent computational research in the life sciences. Genome Biol. 11, R86.   Gorovsky, M A., Hattman, S., and Pleger, G. L. (1973). (6 N)methyl adenine in the nuclear DNA of a eucaryote,  Tetrahymena  pyriformis. J. Cell Biol. 56, 697-701.   Gottschling, D. E., and Cech, T. R. (1984). Chromatin structure of the molecular ends of  Oxytricha  macronuclear DNA: phased nucleosomes and a telomeric complex. Cell 38, 501-510.   Greer, E. L., Blanco, M. A., Gu, L., Sendinc, E., Liu, J., Aristizabal-Corrales, D., Hsu, C.-H., Aravind, L., He, C., and Shi, Y. (2015). DNA Methylation on N6-Adenine in  C. elegans . Cell 161, 868-878.   Haberle, V., Forrest, A. R. R., Hayashizaki, Y., Carninci, P., and Lenhard, B. (2015). CAGEr: precise TSS data retrieval and high-resolution promoterome mining for integrative analyses. Nucleic Acids Res. 43, e51.   Hattman, S., Kenny, C., Berger, L, and Pratt, K. (1978). Comparative study of DNA methylation in three unicellular eucaryotes. J. Bacteriol. 135, 1156-1157.   Horton, J. R., Liebert, K., Bekes, M., Jeltsch, A., and Cheng, X. (2006). Structure and substrate recognition of the  Escherichia coli  DNA adenine methyltransferase. J. Mol. Biol. 358, 559-570.   Huang, Y., Niu, B., Gao, Y., Fu, L., and U, W. (2010). CD-HIT Suite: a web server for clustering and comparing biological sequences. Bioinformatics 26, 680-682.   Huang, J., Dong, X., Gong, Z., Qin, L-Y., Yang, S., Zhu, Y.-L, Wang, X., Zhang, D., Zou, T., Yin, P., et al. (2019). Solution structure of the RNA recognition domain of METTL3-METTL14 N6-methyladenosine methyltransferase. Protein Cell 10, 272-284.   Huff, J. T., and Zilberman, D. (2014). Dnmt1-independent CG methylation contributes to nucleosome positioning in diverse eukaryotes. Cell 156, 1286-1297.   Ito, T., Bulger, M., Pazin, M. J., Kobayashi, R., and Kadonaga, J. T. (1997). ACF, an ISWI-containing and ATP-utilizing chromatin assembly and remodeling factor. Cell 90, 145-155.   Iyer, L M., Zhang, D., and Aravind, L (2016). Adenine methylation in eukaryotes: Apprehending the complex evolutionary history and functional potential of an epigenetic modification. BioEssays 38, 27-40.   Karrer, K. M., and VanNuland, T. A. (1999). Nucleosome positioning is independent of histone H1 in vivo. J. Biol. Chem. 274, 33020-33024.   Katoh, K., Rozewicki, J., and Yamada, K. D. (2017). MAFFT online service: multiple sequence alignment, interactive sequence choice and visualization. Brief. Bioinform.   Kharchenko, P. V., Tolstorukov, M. Y., and Park, P. J. (2008). Design and analysis of ChIP-seq experiments for DNA-binding proteins. Nat. Biotechnol. 26, 1351-1359.   Khurana, J. S., Wang, X., Chen, X., Perlman, D. H., and Landweber, L F. (2014). Transcription-independent functions of an RNA polymerase II subunit, Rpb2, during genome rearrangement in the ciliate,  Oxytricha trifallax . Genetics 197, 839-849.   Khurana, J. S., Clay, D. M., Moreira, S., Wang, X., and Landweber, L. F. (2018). Small RNA-mediated regulation of DNA dosage in the ciliate  Oxytricha . RNA 24, 18-29.   Koziol, M. J., Bradshaw, C. R., Allen, G. E., Costa, A. S. H., Frezza, C., and Gurdon, J. B. (2016). Identification of methylated deoxyadenosines in vertebrates reveals diversity in DNA modifications. Nat. Struct. Mol. Biol. 23, 24-30.   Kuraku, S., Zmasek, C. M., Nishimura, O., and Katoh, K. (2013). aLeaves facilitates on-demand exploration of metazoan gene family trees on MAFFT sequence alignment server with enhanced interactivity. Nucleic Acids Res. 41, W22-W28.   Lai, W. K. M., and Pugh, B. F. (2017). Understanding nucleosome dynamics and their links to gene expression and DNA replication. Nat. Rev. Mol. Cell Biol. 18, 548-562.   Langmead, B., and Salzberg, S. L. (2012). Fast gapped-read alignment with Bowtie 2. Nat. Methods 9, 357-359.   Laughlin, T. J., Henry, J. M., Phares, E. F., Long, M. V., and Olins, D. E. (1983). Methods for the Large-Scale Cultivation of an  Oxytricha  (Ciliophora: Hypotrichide). J. Protozool. 30, 63-64.   Lauth, M. R., Spear, B. B., Neumann, J., and Prescott, D. M. (1976). DNA of ciliated protozoa: DNA sequence diminution during macronuclear development of  Oxytricha . Cell 7, 67-74.   Lawn, R. M., Neumann, J. M., Herrick, G., and Prescott, D. M. (1978). The genesize DNA molecules in  Oxytricha . Cold Spring Harb. Symp. Quant. Biol. 42, 483-492.   Liang, Z., Shen, L., Cui, X., Bao, S., Geng, Y., Yu, G., Liang, F., Xie, S., Lu, T., Gu, X., and Yu, H. (2018). DNA N6-Adenine Methylation in  Arabidopsis thaliana . Dev. Cell 45, 406-416.   Lieleg, C., Ketterer, P., Nuebler, J., Ludwigsen, J., Gerland, U., Dietz, H., Mueller-Planitz, F., and Korber, P. (2015). Nucleosome spacing generated by ISWI and CHD1 remodelers is constant regardless of nucleosome density. Mol. Cell. Biol. 35, 1588-1605.   Liu, Y., Tavema, S. D., Muratore, T. L, Shabanowitz, J., Hunt, D. F., and Allis, C. D. (2007). RNAi-dependent H3K27 methylation is required for heterochromatin formation and DNA elimination in  Tetrahymena . Genes Dev. 21, 1530-1545.   Liu, J., Yue, Y., Han, D., Wang, X., Fu, Y., Zhang, L, Jia, G., Yu, M., Lu, Z., Deng, X., et al. (2014). A METTL3-METTL14 complex mediates mammalian nuclear RNA N6-adenosine methylation. Nat. Chem. Biol. 10, 93-95.   Liu, J., Zhu, Y., Luo, G.-Z., Wang, X., Yue, Y., Wang, X., Zong, X., Chen, K., Yin, H., Fu, Y., et al. (2016). Abundant DNA 6 mA methylation during early embryogenesis of zebrafish and pig. Nat. Commun. 7,13052.   Livak, K. J., and Schmittgen, T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) Method. Methods 25, 402-408.   Lugar, K., Rcchatcincr, T. J., and Richmond, T. J. (1900). Proparation of nucleosome core particle from recombinant histones. Methods Enzymol. 304, 3-19.   Luo, G.-Z., Blanco, M. A., Greer, E. L., He, C., and Shi, Y. (2015). DNA N(6)-methyladenine: a new epigenetic mark in eukaryotes? Nat. Rev. Mol. Cell Biol. 16, 705-710.   Luo, G.-Z., Hao, Z., Luo, L, Shen, M., Sparvoli, D., Zheng, Y., Zhang, Z., Weng, X., Chen, K., Cui, Q., et al. (2018). N6-methyldeoxyadenosine directs nucleosome positioning in  Tetrahymena  DNA. Genome Biol. 19, 200.   Mavrich, T. N., loshikhes, I. P., Venters, B. J., Jiang, C., Tomsho, L P., Qi, J., Schuster, S. C., Albert, I., and Pugh, B. F. (2008). A barrier nucleosome model for statistical positioning of nucleosomes throughout the yeast genome. Genome Res. 18, 1073-1083.   Miao, W., Xiong, J., Bowen, J., Wang, W., Liu, Y., Braguinets, O., Grigull, J., Pearlman, R. E., Orias, E., and Gorovsky, M A. (2009). Microarray analyses of gene expression during the  Tetrahymena thermophila  life cycle. PLoS ONE 4,e4429.   Miller, M A., Pfeiffer, W., and Schwartz, T. (2010). Creating the CIPRES Science Gateway for Inference of Large Phylogenetic Trees. Proceedings of the Gateway Computing Environments Workshop (GCE), 14 Nov. 2010, New Orleans, La. pp. 1-8.   Mondo, S. J., Dannebaum, R. O., Kuo, R. C., Louie, K. B., Bewick, A. J., LaButti, K., Haridas, S., Kuo, A., Salamov, A., Ahrendt, S. R., et al. (2017). Widespread adenine N6-methylation of active genes in fungi. Nat. Genet. 49, 964-968.   Mortazavi, A., Williams, B. A., McCue, K., Schaeffer, L., and Wold, B. (2008). Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat. Methods 5, 621-628.   M. M., Fierz, B., Bittova, L., Liszczak, G., and Muir, T. W. (2016). A two-state activation mechanism controls the histone methyltransferase Suv39h1. Nat. Chem. Biol. 12, 188-193.   Murray, i.A., Morgan, R. D., Luyten, Y., Fomenkov, A., Correa, i.R., jr., Dai, Allaw, M. B., Zhang, X., Cheng, X., and Roberts, R. J. (2018). The non-specific adenine DNA methyltransferase M.EcoGll. Nucleic Acids Res. 46, 840-848.   Nesvizhskii, A. I., Keller, A., Kolker, E., and Aebersold, R. (2003). A statistical model for identifying proteins by tandem mass spectrometry. Anal. Chem. 75, 4646-4658.   Nowacki, M., Vijayan, V., Zhou, Y., Schotanus, K., Doak, T. G., and Landweber, L. F. (2008). RNA-mediated epigenetic programming of a genome-rearrangement pathway. Nature 451, 153-158.   Pendleton, K. E., Chen, B., Liu, K., Hunter, 0. V., Xie, Y., Tu, B. P., and Conrad, N. K. (2017). The U6 snRNA m6A Methyltransferase METTL16 Regulates SAM Synthetase Intron Retention. Cell 169, 824-835.   Pratt, K., and Hattman, S. (1981). Deoxyribonucleic acid methylation and chromatin organization in  Tetrahymena thermophila . Mol. Cell. Biol. 1, 600-608.   Prescott, D. M. (1994). The DNA of ciliated protozoa. Microbiol. Rev. 58, 233-267.   Rae, P. M., and Spear, B. B. (1978). Macronuclear DNA of the hypotrichous ciliate  Oxytricha fallax . Proc. Natl. Acad. Sci. USA 75, 4992-4996.   Rappsilber, J., Mann, M., and Ishihama, Y. (2007). Protocol for micro-purification, enrichment, pre-fractionation and storage of peptides for proteomics using StageTips. Nat. Protoc. 2, 1896-1906.   Schaffer, A. A., Aravind, L, Madden, T. L, Shavirin, S., Spouge, J. L., Wolf, Y. I., Koonin, E. V., and Altschul, S. F. (2001). Improving the accuracy of PSI-BLAST protein database searches with composition-based statistics and other refinements. Nucleic Acids Res. 29, 2994-3005.   Schiffers, S., Ebert, C., Rahimoff, R., Kosmatchev, O., Steinbacher, J., Bohne, A.-V., Spada, F., Michalakis, S., Nickelsen, J., Mailer, M., and Carell, T. (2017). Quantitative LC-MS Provides No Evidence for m6 dA or m4 dC in the Genome of Mouse Embryonic Stem Cells and Tissues. Angew. Chem. Int. Ed. Engl. 56, 11268-11271.   Sledz, P., and Jinek, M. (2016). Structural insights into the molecular mechanism of the m(6)A writer complex. eLife 5. Published online Sep. 14, 2016. https://doi.org/10.7554/eLife.18434.   Strahl, B. D., Ohba, R., Cook, R. G., and Allis, C. D. (1999). Methylation of histone H3 at lysine 4 is highly conserved and correlates with transcriptionally active nuclei in  Tetrahymena . Proc. Natl. Acad. Sci. USA 96, 14967-14972.   Struhl, K., and Segal, E. (2013). Determinants of nucleosome positioning. Nat. Struct. Mol. Biol. 20, 267-273.   Swart, E. G., Bracht, J. R., Magrini, V., Minx, P., Chen, X., Zhou, Y., Khurana, J. S., Goldman, A D., Nowacki, M., Schotanus, K., et al. (2013). The  Oxytricha trifallax  macronuclear genome: a complex eukaryotic genome with 16,000 tiny chromosomes. PLoS Biol. 11, e1001473.   Tavema, S. D., Coyne, R. S., and Allis, C. D. (2002). Methylation of histone h3 at lysine 9 targets programmed DNA elimination in  tetrahymena . Cell 110, 701-711.   Wada, R. K., and Spear, B. B. (1980). Nucleosomal organization of macronuclear chromatin in  Oxytricha fallax . Cell Differ. 9, 261-268.   Wang, P., Doxtader, K. A., and Nam, Y. (2016a). Structural Basis for Cooperative Function of Mettl3 and Mettl14 Methyltransferases. Mol. Cell 63, 306-317.   Wang, X., Feng, J., Xue, Y., Guan, Z., Zhang, D., Liu, Z., Gong, Z., Wang, Q., Huang, J., Tang, C., et al. (2016b). Structural basis of N(6)-adenosine methylation by the METTL3-METTL14 complex. Nature 534, 575-578.   Wang, Y., Chen, X., Sheng, Y., Liu, Y., and Gao, S. (2017). N6-adenine DNA methylation is associated with the linker DNA of H2A2-containing well-positioned nucleosomes in Pol II-transcribed genes in  Tetrahymena . Nucleic Acids Res. 45, 11594-11606.   Warda, A. S., Kretschmer, J., Heckert, P., Lenz, C., Urlaub, H., Hobartner, C., Sloan, K. E., and Bohnsack, M. T. (2017). Human METTL16 is a N6-methyladenosine (m6A) methyltransferase that targets pre-mRNAs and various noncoding RNAs. EMBO Rep. 18, 2004-2014.   Wei, Y., Mizzen, C. A., Cook, R. G., Gorovsky, M. A., and Allis, C. D. (1998). Phosphorylation of histone H3 at serine 10 is correlated with chromosome condensation during mitosis and meiosis in  Tetrahymena . Proc. Natl. Acad. Sci. USA 95, 7480-7484.   Wu, T. P., Wang, T., Seetin, M. G., Lai, Y., Zhu, S., Lin, K., Liu, Y., Byrum, S. D., Mackintosh, S. G., Zhong, M., et al. (2016). DNA methylation on N(6)-adenine in mammalian embryonic stem cells. Nature 532, 329-333.   Xiao, R., and Moore, D. D. (2011). Dam IP: Using Mutant DNA Adenine Methyltransferase to Study DNA-Protein Interactions In Vivo. Curr. Protoc. Mol. Biol. 21. https://doi.org/10.1002/0471142727.mb2121s94.   Xiao, C.-L., Zhu, S., He, M., Chen, D., Zhang, Q., Chen, Y., Yu, G., Liu, J., Xie, S.-Q., Luo, F., et al. (2018). N6-Methyladenine DNA Modification in the Human Genome. Mol. Cell 71, 306-318.   Xiong, J., Lu, X., Lu, Y., Zeng, H., Yuan, D., Feng, L, Chang, Y., Bowen, J., Gorovsky, M., Fu, C., and Miao, W. (2011).  Tetrahymena  Gene Expression Database (TGED): a resource of microarray data and co-expression analyses for  Tetrahymena . Sci. China Life Sci. 54, 65-67.   Xiong, J., Lu, X., Zhou, Z., Chang, Y., Yuan, D., Tian, M., Zhou, Z., Wang, L, Fu, C., Orias, E., and Miao, W. (2012). Transcriptome analysis of the model protozoan,  Tetrahymena thermophila , using Deep RNA sequencing. PLoS ONE 7, e30630.   Yao, B., Cheng, Y., Wang, Z., Li, Y., Chen, L, Huang, L., Zhang, W., Chen, D., Wu, H., Tang, B., and Jin, P. (2017). DNA N6-methyladenine is dynamically regulated in the mouse brain following environmental stress. Nat. Commun. 8, 1122.   Yao, B., Li, Y., Wang, Z., Chen, L., Poidevin, M., Zhang, C., Lin, L, Wang, F., Bao, H., Jiao, B., et al. (2018). Active N 6-Methyladenine Demethylation by DMAD Regulates Gene Expression by Coordinating with Polycomb Protein in Neurons. Mol. Cell 71, 848-857.   Yerlici, V. T., and Landweber, L. F. (2014). Programmed Genome Rearrangements in the Ciliate  Oxytricha . Microbiol. Spectr. 2. Published online December 2014. 10.1128/microbiolspec.MDNA3-0025-2014.   Zhang, Z., and Pugh, B. F. (2011). High-resolution genome-wide mapping of the primary structure of chromatin. Cell 144, 175-186.   Zhang, G., Huang, H., Liu, D., Cheng, Y., Liu, X., Zhang, W., Yin, R., Zhang, D., Zhang, P., Liu, J., et al. (2015). N6-methyladenine DNA modification in  Drosophila . Cell 161, 893-906.   Zhou, C., Wang, C., Liu, H., Zhou, Q., Liu, Q., Guo, Y., Peng, T., Song, J., Zhang, J.,   Chen, L., et al. (2018). Identification and analysis of adenine N6-methylation sites in the rice genome. Nat. Plants 4, 554-563.   

     The embodiments described in this disclosure can be combined in various ways. Any aspect or feature that is described for one embodiment can be incorporated into any other embodiment mentioned in this disclosure. While various novel features of the inventive principles have been shown, described and pointed out as applied to particular embodiments thereof, it should be understood that various omissions and substitutions and changes may be made by those skilled in the art without departing from the spirit of this disclosure. Those skilled in the art will appreciate that the inventive principles can be practiced in other than the described embodiments, which are presented for purposes of illustration and not limitation.