Patent Publication Number: US-2003226179-A1

Title: Plant aminoacyl-tRNA synthetases

Description:
[0001] This application is a divisional of U.S. patent application Ser. No. 09/436699, filed Nov. 4, 1999, which is hereby incorporated herein by reference in its entirety which claims the benefit of U.S. Provisional Application No. 60/107,276, filed Nov. 5, 1998. 
    
    
     
       FIELD OF THE INVENTION  
       [0002] This invention is in the field of plant molecular biology. More specifically, this invention pertains to nucleic acid fragments encoding aminoacyl-tRNA synthetases in plants and seeds.  
       BACKGROUND OF THE INVENTION  
       [0003] Aminoacyl-tRNA Synthetases (AARS) are enzymes that charge (acylate) tRNAs with amino acids. These charged aminoacyl tRNAs then participate in mRNA translation and protein synthesis. The AARS show high specificity for charging a specific tRNA with the appropriate amino acid, for example valyl-tRNA with valine by valyl-tRNA synthetase or tryptophanyl-tRNA with tryptophan by tryptophanyl-tRNA synthetase. In general, per organism there are is at least one AARS for each of the twenty amino acids. There are exceptions however. AARS are ancient enzymes, having functioned in translation since early life evolution. Some have speculated that the earliest aminoacyl-tRNA synthetases were mRNAs, not proteins, with the proteinaceous AARS described here emerging later (Neidhardt et al., (1975)  Annu. Rev. Microbiol.  29:215-250). AARS are structurally diverse, although AARSs for some amino acids are more closely related than for others. AARSs are generally divided into two classes, class I and class II, based on structural similarity and amino acid preferences (Eriani et al., (1990)  Nature  347:203-206).  
       [0004] Plants like all other cellular organisms have aminoacyl-tRNA synthetases. However, a full description of the plant ‘complement’ of aminoacyl-tRNA synthetases has not yet been described. Full-length cDNA, genomic clones, and EST sequences for a variety of plant aminoacyl-tRNA synthetases are known. However, several anticipated aminoacyl-tRNA synthetases have not been discovered.  
       [0005] Because of the central role of protein synthesis in life, any agent that inhibits or disrupts this activity is likely to be toxic. Aminoacyl-tRNA synthetases play a critical role in protein translation by linking genetic nucleic acid information to protein synthesis. Aminoacyl-tRNA synthetases perform this role by “reading” the identity of the different tRNAs and acylating them with the correct cognate amino acid. A large volume of research over several decades has been focused on identifying inhibitors of this process. Inhibitors of aminoacyl-tRNA synthetases have been found to be cytotoxic due to their inhibition of protein synthesis. As such they therefore could be used as herbicides or in aminoacyl-tRNA synthetase selectable marker systems (Lloyd et al., (1995)  Nucleic Acid Research  23(15):2882-2892). The genes disclosed herein can serve as the basis for testing whether the encoded aminoacyl-tRNA synthetases are sensitive to known inhibitors or other chemicals.  
       [0006] Biochemical processes are often compartmentalized in regions of cells, such as mitochondria, plastids, and lysosomes. These organelles are key sites for many biochemical pathways. Bioengineering of these processes may require targeting protein products to specific organelles. One method to accomplish this involves the addition of an N-terminal prosequence (transit peptide) that directs protein entry into a specific organelle(s). Upon or shortly after transport into the organelle the transit peptide is usually proteolytically removed, and the mature protein is then able to function.  
       [0007] A few plant transit peptides have been shown empirically to be capable of directing fused proteins into specific organelles. However this ability appears to depend upon the structure of the protein being imported and to date it is impossible to predict whether a protein will be imported into an organelle with a given transit peptide. As such, it is advantageous to have a diversity of potential transit peptides from which the most efficient candidate can be chosen to target a protein of interest to an organelle. A number of plant transit peptides are known which direct mature proteins to mitochondria or chloroplast organelles. These transit peptides are diverse in structure (length and amino acid sequence) and there is no strong consensus sequence identifying them. In addition, there is no obvious clear relationship between chloroplast targeting and mitochondrial targeting transit sequences. This invention describes a number of chloroplast-targeting and mitochondria-targeting transit peptides for (maize) aminoacyl-tRNA synthetases. These sequences will find utility in directing both aminoacyl-tRNA synthetase and other proteins into these organelles.  
       [0008] Accordingly, the availability of nucleic acid sequences encoding all or a portion of these enzymes would facilitate studies to better understand protein synthesis in plants, provide genetic tools for the manipulation of gene expression, protein targeting to specific organelles and provide possible targets for herbicides.  
       SUMMARY OF THE INVENTION  
       [0009] The present invention relates to isolated polynucleotides comprising a nucleotide sequence encoding a first polypeptide of at least 116 amino acids that has at least 85% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of a rice seryl-tRNA synthetase polypeptide of SEQ ID NO:16, a soybean seryl-tRNA synthetase polypeptide of SEQ ID NO:18, or a wheat seryl-tRNA synthetase polypeptide of SEQ ID NO:20. The present invention also relates to an isolated polynucleotide comprising the complement of the nucleotide sequences described above.  
       [0010] The present invention also relates to isolated polynucleotides comprising a nucleotide sequence encoding a second polypeptide of at least 483 amino acids that has at least 80% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of a corn threonyl-tRNA synthetase polypeptide of SEQ ID NO:22, a rice threonyl-tRNA synthetase polypeptide of SEQ ID NO:24, a soybean threonyl-tRNA synthetase polypeptide of SEQ ID NO:26, or a wheat threonyl-tRNA synthetase polypeptide of SEQ ID NO:28. The present invention also relates to an isolated polynucleotide comprising the complement of the nucleotide sequences described above.  
       [0011] The present invention relates to an isolated polynucleotide comprising a nucleotide sequence encoding a first polypeptide of at least 30 amino acids that has at least 70% identity based on the Clustal method of alignment when compared to a polypeptide selected from the group consisting of SEQ ID NOs:2, 4, 6, 8, 10, 12 and 14.  
       [0012] It is preferred that the isolated polynucleotides of the claimed invention consists of a nucleic acid sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25 and 27 that codes for the polypeptide selected from the group consisting of SEQ ID NOs:2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26 and 28. The present invention also relates to an isolated polynucleotide comprising a nucleotide sequences of at least one of 40 (preferably at least one of 30) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25 and 27 and the complement of such nucleotide sequences.  
       [0013] The present invention relates to a chimeric gene comprising an isolated polynucleotide of the present invention operably linked to suitable regulatory sequences.  
       [0014] The present invention also relates to the identification of transit peptides associated with aminoacyl-tRNA synthetases of the instant invention and the use of those transit peptides to target aminoacyl-tRNA synthetases and other operably linked proteins to specific organelles within plant cells. Transit peptide amino acid sequences are located just upstream of the mature aminoacyl-tRNA synthetase polypeptide sequences disclosed in the instant invention.  
       [0015] The present invention relates to an isolated host cell comprising a chimeric gene of the present invention or an isolated polynucleotide of the present invention. The host cell may be eukaryotic, such as a yeast or a plant cell, or prokaryotic, such as a bacterial cell. The present invention also relates to a virus, preferably a baculovirus, comprising an isolated polynucleotide of the present invention or a chimeric gene of the present invention.  
       [0016] The present invention relates to a process for producing an isolated host cell comprising a chimeric gene of the present invention or an isolated polynucleotide of the present invention, the process comprising either transforming or transfecting an isolated compatible host cell with a chimeric gene or isolated polynucleotide of the present invention.  
       [0017] The present invention relates to a seryl-tRNA synthetase polypeptide of at least 116 amino acids comprising at least 85% homology based on the Clustal method of alignment compared to a polypeptide selected from the group consisting of SEQ ID NOs:16, 18 and 20.  
       [0018] The present invention alos also relates to a threonyl-tRNA synthetase polypeptide of at least 483 amino acids comprising at least 80% homology based on the Clustal method of alignment compared to a polypeptide selected from the group consisting of SEQ ID NOs:20, 22, 24, 26 and 28.  
       [0019] The present invention relates to a method of selecting an isolated polynucleotide that affects the level of expression of a seryl-tRNA synthetase or threonyl-tRNA synthetase polypeptide in a host cell, preferably a plant cell, the method comprising the steps of:  
       [0020] constructing an isolated polynucleotide of the present invention or an isolated chimeric gene of the present invention;  
       [0021] introducing the isolated polynucleotide or the isolated chimeric gene into a host cell;  
       [0022] measuring the level a seryl-tRNA synthetase or threonyl-tRNA synthetase polypeptide in the host cell containing the isolated polynucleotide; and  
       [0023] comparing the level of a seryl-tRNA synthetase or threonyl-tRNA synthetase polypeptide in the host cell containing the isolated polynucleotide with the level of a seryl-tRNA synthetase or threonyl-tRNA synthetase polypeptide in a host cell that does not contain the isolated polynucleotide.  
       [0024] The present invention relates to a method of obtaining a nucleic acid fragment encoding a substantial portion of a seryl-tRNA synthetase or threonyl-tRNA synthetase polypeptide gene, preferably a plant seryl-tRNA synthetase or threonyl-tRNA synthetase polypeptide gene, comprising the steps of: synthesizing an oligonucleotide primer comprising a nucleotide sequence of at least one of 40 (preferably at least one of 30) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25 and 27 and the complement of such nucleotide sequences; and amplifying a nucleic acid fragment (preferably a cDNA inserted in a cloning vector) using the oligonucleotide primer. The amplified nucleic acid fragment preferably will encode a portion of a seryl-tRNA synthetase or threonyl-tRNA synthetase amino acid sequence.  
       [0025] The present invention also relates to a method of obtaining a nucleic acid fragment encoding all or a substantial portion of the amino acid sequence encoding a seryl-tRNA synthetase or threonyl-tRNA synthetase polypeptide comprising the steps of: probing a cDNA or genomic library with an isolated polynucleotide of the present invention; identifying a DNA clone that hybridizes with an isolated polynucleotide of the present invention; isolating the identified DNA clone; and sequencing the cDNA or genomic fragment that comprises the isolated DNA clone.  
       [0026] A further embodiment of the instant invention is a method for evaluating at least one compound for its ability to inhibit the activity of a seryl-tRNA synthetase or threonyl-tRNA synthetase, the method comprising the steps of: (a) transforming a host cell with a chimeric gene comprising a nucleic acid fragment encoding an aminoacyl-tRNA synthetase polypeptide, operably linked to suitable regulatory sequences; (b) growing the transformed host cell under conditions that are suitable for expression of the chimeric gene wherein expression of the chimeric gene results in production of an aminoacyl-tRNA synthetase polypeptide in the transformed host cell; (c) optionally purifying the aminoacyl-tRNA synthetase expressed by the transformed host cell; (d) treating the amino-acyl-tRNA synthetase with a compound to be tested; and (e) comparing the activity of the aminoacyl-tRNA synthetase that has been treated with a test compound to the activity of an untreated aminoacyl-tRNA synthetase, thereby selecting compounds with potential for inhibitory activity.  
       [0027] The present invention relates to a composition comprising an isolated polynucleotide or polypeptide of the present invention.  
       [0028] The present invention relates to an expression cassette comprising an isolated polynucleotide of the present invention.  
       [0029] The present invention relates to a method for positive selection of a transformed cell comprising:  
       [0030] (a) transforming a plant cell, preferably a monocot such as corn, with a chimeric gene of the present invention or an expression cassette of the present invention; and  
       [0031] (b) growing the transformed plant under conditions allowing expression of the polynucleotide in an amount sufficient to complement an amiono acyl-tRNA synthetase.  
       [0032] As used herein, the following terms shall apply:  
       [0033] “Aminoacyl-tRNA synthetase” refers to seryl-tRNA synthetase and/or threonyl-tRNA synthetase.  
       BRIEF DESCRIPTION OF THE SEQUENCE DESCRIPTIONS  
       [0034] The invention can be more fully understood from the following detailed description and the accompanying Sequence Listing which form a part of this application.  
       [0035] Table 1 lists the polypeptides that are described herein, the designation of the cDNA clones that comprise the nucleic acid fragments encoding polypeptides representing all or a substantial portion of these polypeptides, and the corresponding identifier (SEQ ID NO:) as used in the attached Sequence Listing. Table 1 also identifies the cDNA clones as individual ESTs (“EST”), the sequences of the entire cDNA inserts comprising the indicated cDNA clones (“FIS”), contigs assembled from two or more ESTs (“Contig”), contigs assembled from an FIS and one or more ESTs (“Contig*”), or sequences encoding the entire protein derived from an FIS, a contig, or an FIS and PCR (“CGS”). Nucleotide sequences, SEQ ID NOs:15, 17, 19, 21, 23, 25 and 27 and amino acid sequences SEQ ID NOs:16, 18, 20, 22, 24, 26 and 28 were determined by further sequence analysis of cDNA clones encoding the amino acid sequences set forth in SEQ ID NOs:2, 4, 6, 8, 10, 12 and 14. Nucleotide SEQ ID NOs:1, 3, 5, 7, 9, 11 and 13 and amino acid SEQ ID NOs:2, 4, 6, 8, 10, 12 and 14 were presented in a U.S. Provisional Application No. 60/107,276, filed Nov. 5, 1998.  
       [0036] The sequence descriptions and Sequence Listing attached hereto comply with the rules governing nucleotide and/or amino acid sequence disclosures in patent applications as set forth in 37 C.F.R. §1.821-1.825.  
               TABLE 1                          Aminoacyl-tRNA Synthetases                         SEQ ID NO:                                     (Nucle-   (Amino       Protein   Clone Designation   otide)   Acid)                                     Seryl-tRNA Synthetase   r1r6.pk0023.e9 (EST)   1   2       Seryl-tRNA Synthetase   s12.pk123.g22 (EST)   3   4       Seryl-tRNA Synthetase   w1m4.pk008.d5 (EST)   5   6       Threonyl-tRNA Synthetase   cr1n.pk0140.b10 (EST)   7   8       Threonyl-tRNA Synthetase   r1r6.pk0084.e1 (EST)   9   10       Threonyl-tRNA Synthetase   srr1c.pk003.k12 (EST)   11   12       Threonyl-tRNA Synthetase   wr1.pk181.e1 (EST)   13   14       Seryl-tRNA Synthetase   r1r6.pk0023.e9 (CGS)   15   16       Seryl-tRNA Synthetase   s12.pk123.g22 (FIS)   17   18       Seryl-tRNA Synthetase   w1m4.pk0018.d5 (CGS)   19   20       Threonyl-tRNA Synthetase   p0017.cespf50r (CGS)   21   22       Threonyl-tRNA Synthetase   res1c.pk005.j13 (FIS) 23   24       Threonyl-tRNA Synthetase   srr1c.pk003.k12 (FIS)   25   26       Threonyl-tRNA_Synthetase   w1m96.pk060.l13 (FIS)   27   28                  
 
       [0037] The Sequence Listing contains the one letter code for nucleotide sequence characters and the three letter codes for amino acids as defined in conformity with the IUPAC-IUBMB standards described in  Nucleic Acids Res.  13:3021-3030 (1985) and in the  Biochemical J.  219 (No. 2):345-373 (1984) which are herein incorporated by reference. The symbols and format used for nucleotide and amino acid sequence data comply with the rules set forth in 37 C.F.R. §1.822.  
       DETAILED DESCRIPTION OF THE INVENTION  
       [0038] In the context of this disclosure, a number of terms shall be utilized. As used herein, a “polynucleotide” is a nucleotide sequence such as a nucleic acid fragment. A polynucleotide may be a polymer of RNA or DNA that is single- or double-stranded, that optionally contains synthetic, non-natural or altered nucleotide bases. A polynucleotide in the form of a polymer of DNA may be comprised of one or more segments of cDNA, genomic DNA, or synthetic DNA. An isolated polynucleotide of the present invention may include at least one of 60 contiguous nucleotides, preferably at least one of 40 contiguous nucleotides, most preferably one of at least 30 contiguous nucleotides, of the nucleic acid sequence of the SEQ ID NOs:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25 and 27.  
       [0039] As used herein, “substantially similar” refers to nucleic acid fragments wherein changes in one or more nucleotide bases results in substitution of one or more amino acids, but do not affect the functional properties of the polypeptide encoded by the nucleotide sequence. “Substantially similar” also refers to nucleic acid fragments wherein changes in one or more nucleotide bases does not affect the ability of the nucleic acid fragment to mediate alteration of gene expression by gene silencing through for example antisense or co-suppression technology. “Substantially similar” also refers to modifications of the nucleic acid fragments of the instant invention such as deletion or insertion of one or more nucleotides that do not substantially affect the functional properties of the resulting transcript vis-à-vis the ability to mediate gene silencing or alteration of the functional properties of the resulting protein molecule. It is therefore understood that the invention encompasses more than the specific exemplary nucleotide or amino acid sequences and includes functional equivalents thereof.  
       [0040] Substantially similar nucleic acid fragments may be selected by screening nucleic acid fragments representing subfragments or modifications of the nucleic acid fragments of the instant invention, wherein one or more nucleotides are substituted, deleted and/or inserted, for their ability to affect the level of the polypeptide encoded by the unmodified nucleic acid fragment in a plant or plant cell. For example, a substantially similar nucleic acid fragment representing at least one of 30 contiguous nucleotides derived from the instant nucleic acid fragment can be constructed and introduced into a plant or plant cell. The level of the polypeptide encoded by the unmodified nucleic acid fragment present in a plant or plant cell exposed to the substantially similar nucleic fragment can then be compared to the level of the polypeptide in a plant or plant cell that is not exposed to the substantially similar nucleic acid fragment.  
       [0041] For example, it is well known in the art that antisense suppression and co-suppression of gene expression may be accomplished using nucleic acid fragments representing less than the entire coding region of a gene, and by nucleic acid fragments that do not share 100% sequence identity with the gene to be suppressed. Moreover, alterations in a nucleic acid fragment which result in the production of a chemically equivalent amino acid at a given site, but do not effect the functional properties of the encoded polypeptide, are well known in the art. Thus, a codon for the amino acid alanine, a hydrophobic amino acid, may be substituted by a codon encoding another less hydrophobic residue, such as glycine, or a more hydrophobic residue, such as valine, leucine, or isoleucine. Similarly, changes which result in substitution of one negatively charged residue for another, such as aspartic acid for glutamic acid, or one positively charged residue for another, such as lysine for arginine, can also be expected to produce a functionally equivalent product. Nucleotide changes which result in alteration of the N-terminal and C-terminal portions of the polypeptide molecule would also not be expected to alter the activity of the polypeptide. Each of the proposed modifications is well within the routine skill in the art, as is determination of retention of biological activity of the encoded products. Consequently, an isolated polynucleotide comprising a nucleotide sequence of at least one of 60 (preferably at least one of 40, most preferably at least one of 30) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25 and 27 and the complement of such nucleotide sequences may be used in methods of selecting an isolated polynucleotide that affects the expression of a polypeptide (such as aminoacyl-tRNA synthetase) in a plant cell. A method of selecting an isolated polynucleotide that affects the level of expression of a polypeptide in a host cell (eukaryotic, such as plant or yeast, prokaryotic such as bacterial, or viral) may comprise the steps of: constructing an isolated polynucleotide of the present invention or an isolated chimeric gene of the present invention; introducing the isolated polynucleotide or the isolated chimeric gene into a host cell; measuring the level of a polypeptide in the host cell containing the isolated polynucleotide; and comparing the level of a polypeptide in the host cell containing the isolated polynucleotide with the level of a polypeptide in a host cell that does not contain the isolated polynucleotide.  
       [0042] Moreover, substantially similar nucleic acid fragments may also be characterized by their ability to hybridize. Estimates of such homology are provided by either DNA-DNA or DNA-RNA hybridization under conditions of stringency as is well understood by those skilled in the art (Hames and Higgins, Eds. (1985) Nucleic Acid Hybridisation, IRL Press, Oxford, U.K.). Stringency conditions can be adjusted to screen for moderately similar fragments, such as homologous sequences from distantly related organisms, to highly similar fragments, such as genes that duplicate functional enzymes from closely related organisms. Post-hybridization washes determine stringency conditions. One set of preferred conditions uses a series of washes starting with 6×SSC, 0.5% SDS at room temperature for 15 min, then repeated with 2×SSC, 0.5% SDS at 45° C. for 30 min, and then repeated twice with 0.2×SSC, 0.5% SDS at 50° C. for 30 min. A more preferred set of stringent conditions uses higher temperatures in which the washes are identical to those above except for the temperature of the final two 30 min washes in 0.2×SSC, 0.5% SDS was increased to 60° C. Another preferred set of highly stringent conditions uses two final washes in 0.1×SSC, 0.1% SDS at 65° C.  
       [0043] Substantially similar nucleic acid fragments of the instant invention may also be characterized by the percent identity of the amino acid sequences that they encode to the amino acid sequences disclosed herein, as determined by algorithms commonly employed by those skilled in this art. Suitable nucleic acid fragments (isolated polynucleotides of the present invention) encode polypeptides that are at least 70% identical, preferably at least 80% identical to the amino acid sequences reported herein. Preferred nucleic acid fragments encode amino acid sequences that are at least 85% identical to the amino acid sequences reported herein. More preferred nucleic acid fragments encode amino acid sequences that are at least 90% identical to the amino acid sequences reported herein. Most preferred are nucleic acid fragments that encode amino acid sequences that are at least 95% identical to the amino acid sequences reported herein. Suitable nucleic acid fragments not only have the above homologies but typically encode a polypeptide having at least 50 amino acids, preferably at least 100 amino acids, more preferably at least 150 amino acids, still more preferably at least 200 amino acids, and most preferably at least 250 amino acids. Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal method of alignment (Higgins and Sharp (1989)  CABIOS.  5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5.  
       [0044] A “substantial portion” of an amino acid or nucleotide sequence comprises an amino acid or a nucleotide sequence that is sufficient to afford putative identification of the protein or gene that the amino acid or nucleotide sequence comprises. Amino acid and nucleotide sequences can be evaluated either manually by one skilled in the art, or by using computer-based sequence comparison and identification tools that employ algorithms such as BLAST (Basic Local Alignment Search Tool; Altschul et al. (1993)  J. Mol. Biol.  215:403-410; see also www.nebi.nlm.nih.gov/BLAST/). In general, a sequence of ten or more contiguous amino acids or thirty or more contiguous nucleotides is necessary in order to putatively identify a polypeptide or nucleic acid sequence as homologous to a known protein or gene. Moreover, with respect to nucleotide sequences, gene-specific oligonucleotide probes comprising 30 or more contiguous nucleotides may be used in sequence-dependent methods of gene identification (e.g., Southern hybridization) and isolation (e.g., in situ hybridization of bacterial colonies or bacteriophage plaques). In addition, short oligonucleotides of 12 or more nucleotides may be used as amplification primers in PCR in order to obtain a particular nucleic acid fragment comprising the primers. Accordingly, a “substantial portion” of a nucleotide sequence comprises a nucleotide sequence that will afford specific identification and/or isolation of a nucleic acid fragment comprising the sequence. The instant specification teaches amino acid and nucleotide sequences encoding polypeptides that comprise one or more particular plant proteins. The skilled artisan, having the benefit of the sequences as reported herein, may now use all or a substantial portion of the disclosed sequences for purposes known to those skilled in this art. Accordingly, the instant invention comprises the complete sequences as reported in the accompanying Sequence Listing, as well as substantial portions of those sequences as defined above.  
       [0045] “Codon degeneracy” refers to divergence in the genetic code permitting variation of the nucleotide sequence without effecting the amino acid sequence of an encoded polypeptide. Accordingly, the instant invention relates to any nucleic acid fragment comprising a nucleotide sequence that encodes all or a substantial portion of the amino acid sequences set forth herein. The skilled artisan is well aware of the “codon-bias” exhibited by a specific host cell in usage of nucleotide codons to specify a given amino acid. Therefore, when synthesizing a nucleic acid fragment for improved expression in a host cell, it is desirable to design the nucleic acid fragment such that its frequency of codon usage approaches the frequency of preferred codon usage of the host cell.  
       [0046] “Synthetic nucleic acid fragments” can be assembled from oligonucleotide building blocks that are chemically synthesized using procedures known to those skilled in the art. These building blocks are ligated and annealed to form larger nucleic acid fragments which may then be enzymatically assembled to construct the entire desired nucleic acid fragment. “Chemically synthesized”, as related to nucleic acid fragment, means that the component nucleotides were assembled in vitro. Manual chemical synthesis of nucleic acid fragments may be accomplished using well established procedures, or automated chemical synthesis can be performed using one of a number of commercially available machines. Accordingly, the nucleic acid fragments can be tailored for optimal gene expression based on optimization of nucleotide sequence to reflect the codon bias of the host cell. The skilled artisan appreciates the likelihood of successful gene expression if codon usage is biased towards those codons favored by the host. Determination of preferred codons can be based on a survey of genes derived from the host cell where sequence information is available.  
       [0047] “Gene” refers to a nucleic acid fragment that expresses a specific protein, including regulatory sequences preceding (5′ non-coding sequences) and following (3′ non-coding sequences) the coding sequence. “Native gene” refers to a gene as found in nature with its own regulatory sequences. “Chimeric gene” refers any gene that is not a native gene, comprising regulatory and coding sequences that are not found together in nature. Accordingly, a chimeric gene may comprise regulatory sequences and coding sequences that are derived from different sources, or regulatory sequences and coding sequences derived from the same source, but arranged in a manner different than that found in nature. “Endogenous gene” refers to a native gene in its natural location in the genome of an organism. A “foreign” gene refers to a gene not normally found in the host organism, but that is introduced into the host organism by gene transfer. Foreign genes can comprise native genes inserted into a non-native organism, or chimeric genes. A “transgene” is a gene that has been introduced into the genome by a transformation procedure.  
       [0048] “Coding sequence” refers to a nucleotide sequence that codes for a specific amino acid sequence. “Regulatory sequences” refer to nucleotide sequences located upstream (5′ non-coding sequences), within, or downstream (3′ non-coding sequences) of a coding sequence, and which influence the transcription, RNA processing or stability, or translation of the associated coding sequence. Regulatory sequences may include promoters, translation leader sequences, introns, and polyadenylation recognition sequences.  
       [0049] “Promoter” refers to a nucleotide sequence capable of controlling the expression of a coding sequence or functional RNA. In general, a coding sequence is located 3′ to a promoter sequence. The promoter sequence consists of proximal and more distal upstream elements, the latter elements often referred to as enhancers. Accordingly, an “enhancer” is a nucleotide sequence which can stimulate promoter activity and may be an innate element of the promoter or a heterologous element inserted to enhance the level or tissue-specificity of a promoter. Promoters may be derived in their entirety from a native gene, or be composed of different elements derived from different promoters found in nature, or even comprise synthetic nucleotide segments. It is understood by those skilled in the art that different promoters may direct the expression of a gene in different tissues or cell types, or at different stages of development, or in response to different environmental conditions. Promoters which cause a nucleic acid fragment to be expressed in most cell types at most times are commonly referred to as “constitutive promoters”. New promoters of various types useful in plant cells are constantly being discovered; numerous examples may be found in the compilation by Okamuro and Goldberg (1989)  Biochemistry of Plants  15:1-82. It is further recognized that since in most cases the exact boundaries of regulatory sequences have not been completely defined, nucleic acid fragments of different lengths may have identical promoter activity.  
       [0050] The “translation leader sequence” refers to a nucleotide sequence located between the promoter sequence of a gene and the coding sequence. The translation leader sequence is present in the fully processed mRNA upstream of the translation start sequence. The translation leader sequence may affect processing of the primary transcript to mRNA, mRNA stability or translation efficiency. Examples of translation leader sequences have been described (Turner and Foster (1995)  Mol. Biotechnol.  3:225-236).  
       [0051] The “3′ non-coding sequences” refer to nucleotide sequences located downstream of a coding sequence and include polyadenylation recognition sequences and other sequences encoding regulatory signals capable of affecting mRNA processing or gene expression. The polyadenylation signal is usually characterized by affecting the addition of polyadenylic acid tracts to the 3′ end of the mRNA precursor. The use of different 3′ non-coding sequences is exemplified by Ingelbrecht et al. (1989)  Plant Cell  1:671-680.  
       [0052] “RNA transcript” refers to the product resulting from RNA polymerase-catalyzed transcription of a DNA sequence. When the RNA transcript is a perfect complementary copy of the DNA sequence, it is referred to as the primary transcript or it may be a RNA sequence derived from post-transcriptional processing of the primary transcript and is referred to as the mature RNA. “Messenger RNA (mRNA)” refers to the RNA that is without introns and that can be translated into polypeptide by the cell. “cDNA” refers to a double-stranded DNA that is complementary to and derived from mRNA. “Sense” RNA refers to an RNA transcript that includes the mRNA and so can be translated into a polypeptide by the cell. “Antisense RNA” refers to an RNA transcript that is complementary to all or part of a target primary transcript or mRNA and that blocks the expression of a target gene (see U.S. Pat. No. 5,107,065, incorporated herein by reference). The complementarity of an antisense RNA may be with any part of the specific nucleotide sequence, i.e., at the 5′ non-coding sequence, 3′ non-coding sequence, introns, or the coding sequence. “Functional RNA” refers to sense RNA, antisense RNA, ribozyme RNA, or other RNA that may not be translated but yet has an effect on cellular processes.  
       [0053] The term “operably linked” refers to the association of two or more nucleic acid fragments on a single nucleic acid fragment so that the function of one is affected by the other. For example, a promoter is operably linked with a coding sequence when it is capable of affecting the expression of that coding sequence (i.e., that the coding sequence is under the transcriptional control of the promoter). Coding sequences can be operably linked to regulatory sequences in sense or antisense orientation.  
       [0054] The term “expression”, as used herein, refers to the transcription and stable accumulation of sense (mRNA) or antisense RNA derived from the nucleic acid fragment of the invention. Expression may also refer to translation of mRNA into a polypeptide. “Antisense inhibition” refers to the production of antisense RNA transcripts capable of suppressing the expression of the target protein. “Overexpression” refers to the production of a gene product in transgenic organisms that exceeds levels of production in normal or non-transformed organisms. “Co-suppression” refers to the production of sense RNA transcripts capable of suppressing the expression of identical or substantially similar foreign or endogenous genes (U.S. Pat. No. 5,231,020, incorporated herein by reference).  
       [0055] “Altered levels” refers to the production of gene product(s) in transgenic organisms in amounts or proportions that differ from that of normal or non-transformed organisms.  
       [0056] “Mature” protein refers to a post-translationally processed polypeptide; i.e., one from which any pre- or propeptides present in the primary translation product have been removed. “Precursor” protein refers to the primary product of translation of mRNA; i.e., with pre- and propeptides still present. Pre- and propeptides may be but are not limited to intracellular localization signals.  
       [0057] A “chloroplast transit peptide” is an amino acid sequence which is translated in conjunction with a protein and directs the protein to the chloroplast or other plastid types present in the cell in which the protein is made. “Chloroplast transit sequence” refers to a nucleotide sequence that encodes a chloroplast transit peptide. A “signal peptide” is an amino acid sequence which is translated in conjunction with a protein and directs the protein to the secretory system (Chrispeels (1991)  Ann. Rev. Plant Phys. Plant Mol. Biol.  42:21-53). If the protein is to be directed to a vacuole, a vacuolar targeting signal (supra) can further be added, or if to the endoplasmic reticulum, an endoplasmic reticulum retention signal (supra) may be added. If the protein is to be directed to the nucleus, any signal peptide present should be removed and instead a nuclear localization signal included (Raikhel (1992)  Plant Phys.  100:1627-1632).  
       [0058] “Transformation” refers to the transfer of a nucleic acid fragment into the genome of a host organism, resulting in genetically stable inheritance. Host organisms containing the transformed nucleic acid fragments are referred to as “transgenic” organisms. Examples of methods of plant transformation include Agrobacterium-mediated transformation (De Blaere et al. (1987)  Meth. Enzymol.  143:277) and particle-accelerated or “gene gun” transformation technology (Klein et al. (1987)  Nature  ( London ) 327:70-73; U.S. Pat. No. 4,945,050, incorporated herein by reference).  
       [0059] Standard recombinant DNA and molecular cloning techniques used herein are well known in the art and are described more fully in Sambrook et al.  Molecular Cloning: A Laboratory Manual;  Cold Spring Harbor Laboratory Press: Cold Spring Harbor, 1989 (hereinafter “Maniatis”).  
       [0060] Nucleic acid fragments encoding at least a portion of several aminoacyl-tRNA synthetases have been isolated and identified by comparison of random plant cDNA sequences to public databases containing nucleotide and protein sequences using the BLAST algorithms well known to those skilled in the art. The nucleic acid fragments of the instant invention may be used to isolate cDNAs and genes encoding homologous proteins from the same or other plant species. Isolation of homologous genes using sequence-dependent protocols is well known in the art. Examples of sequence-dependent protocols include, but are not limited to, methods of nucleic acid hybridization, and methods of DNA and RNA amplification as exemplified by various uses of nucleic acid amplification technologies (e.g., polymerase chain reaction, ligase chain reaction).  
       [0061] For example, genes encoding other seryl-tRNA synthetase or threonyl-tRNA synthetase polypeptides, either as cDNAs or genomic DNAs, could be isolated directly by using all or a portion of the instant nucleic acid fragments as DNA hybridization probes to screen libraries from any desired plant employing methodology well known to those skilled in the art. Specific oligonucleotide probes based upon the instant nucleic acid sequences can be designed and synthesized by methods known in the art (Maniatis). Moreover, the entire sequences can be used directly to synthesize DNA probes by methods known to the skilled artisan such as random primer DNA labeling, nick translation, or end-labeling techniques, or RNA probes using available in vitro transcription systems. In addition, specific primers can be designed and used to amplify a part or all of the instant sequences. The resulting amplification products can be labeled directly during amplification reactions or labeled after amplification reactions, and used as probes to isolate full length cDNA or genomic fragments under conditions of appropriate stringency.  
       [0062] In addition, two short segments of the instant nucleic acid fragments may be used in polymerase chain reaction protocols to amplify longer nucleic acid fragments encoding homologous genes from DNA or RNA. The polymerase chain reaction may also be performed on a library of cloned nucleic acid fragments wherein the sequence of one primer is derived from the instant nucleic acid fragments, and the sequence of the other primer takes advantage of the presence of the polyadenylic acid tracts to the 3′ end of the mRNA precursor encoding plant genes. Alternatively, the second primer sequence may be based upon sequences derived from the cloning vector. For example, the skilled artisan can follow the RACE protocol (Frohman et al. (1988)  Proc. Natl. Acad. Sci. USA  85:8998-9002) to generate cDNAs by using PCR to amplify copies of the region between a single point in the transcript and the 3′ or 5′ end. Primers oriented in the 3′ and 5′ directions can be designed from the instant sequences. Using commercially available 3′ RACE or 5′ RACE systems (BRL), specific 3′ or 5′ cDNA fragments can be isolated (Ohara et al. (1989)  Proc. Natl. Acad. Sci. USA  86:5673-5677; Loh et al. (1989)  Science  243:217-220). Products generated by the 3′ and 5′ RACE procedures can be combined to generate full-length cDNAs (Frohman and Martin (1989)  Techniques  1:165). Consequently, a polynucleotide comprising a nucleotide sequence of at least one of 60 (preferably one of at least 40, most preferably one of at least 30) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25 and 27 and the complement of such nucleotide sequences may be used in such methods to obtain a nucleic acid fragment encoding a substantial portion of an amino acid sequence of a polypeptide. The present invention relates to a method of obtaining a nucleic acid fragment encoding a substantial portion of a polypeptide of a gene (such as seryl-tRNA synthetase or threonyl-tRNA synthetase polypeptide) preferably a substantial portion of a plant polypeptide of a gene, comprising the steps of: synthesizing an oligonucleotide primer comprising a nucleotide sequence of at least one of 60 (preferably at least one of 40, most preferably at least one of 30) contiguous nucleotides derived from a nucleotide sequence selected from the group consisting of SEQ ID NOs:1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 23, 25 and 27 and the complement of such nucleotide sequences; and amplifying a nucleic acid fragment (preferably a cDNA inserted in a cloning vector) using the oligonucleotide primer. The amplified nucleic acid fragment preferably will encode a portion of a polypeptide (such as aminoacyl-tRNA synthetase polypeptide).  
       [0063] Availability of the instant nucleotide and deduced amino acid sequences facilitates immunological screening of cDNA expression libraries. Synthetic peptides representing portions of the instant amino acid sequences may be synthesized. These peptides can be used to immunize animals to produce polyclonal or monoclonal antibodies with specificity for peptides or proteins comprising the amino acid sequences. These antibodies can be then be used to screen cDNA expression libraries to isolate full-length cDNA clones of interest (Lerner (1984)  Adv. Immunol.  36:1-34; Maniatis).  
       [0064] The nucleic acid fragments of the instant invention may be used to create transgenic plants in which the disclosed polypeptides are present at higher or lower levels than normal or in cell types or developmental stages in which they are not normally found. This would have the effect of altering the level of aminoacyl-tRNA synthetase in those cells.  
       [0065] Overexpression of the proteins of the instant invention may be accomplished by first constructing a chimeric gene in which the coding region is operably linked to a promoter capable of directing expression of a gene in the desired tissues at the desired stage of development. For reasons of convenience, the chimeric gene may comprise promoter sequences and translation leader sequences derived from the same genes. 3′ Non-coding sequences encoding transcription termination signals may also be provided. The instant chimeric gene may also comprise one or more introns in order to facilitate gene expression.  
       [0066] Plasmid vectors comprising the instant chimeric gene can then be constructed. The choice of plasmid vector is dependent upon the method that will be used to transform host plants. The skilled artisan is well aware of the genetic elements that must be present on the plasmid vector in order to successfully transform, select and propagate host cells containing the chimeric gene. The skilled artisan will also recognize that different independent transformation events will result in different levels and patterns of expression (Jones et al. (1985)  EMBO J.  4:2411-2418; De Almeida et al. (1989)  Mol. Gen. Genetics  218:78-86), and thus that multiple events must be screened in order to obtain lines displaying the desired expression level and pattern. Such screening may be accomplished by Southern analysis of DNA, Northern analysis of mRNA expression, Western analysis of protein expression, or phenotypic analysis.  
       [0067] For some applications it may be useful to direct the instant polypeptides to different cellular compartments, or to facilitate its secretion from the cell. It is thus envisioned that the chimeric gene described above may be further supplemented by altering the coding sequence to encode the instant polypeptides with appropriate intracellular targeting sequences such as transit sequences (Keegstra (1989)  Cell  56:247-253), signal sequences or sequences encoding endoplasmic reticulum localization (Chrispeels (1991)  Ann. Rev. Plant Phys. Plant Mol. Biol.  42:21-53), or nuclear localization signals (Raikhel (1992)  Plant Phys.  100:1627-1632) added and/or with targeting sequences that are already present removed. While the references cited give examples of each of these, the list is not exhaustive and more targeting signals of utility may be discovered in the future.  
       [0068] It may also be desirable to reduce or eliminate expression of genes encoding the instant polypeptides in plants for some applications. In order to accomplish this, a chimeric gene designed for co-suppression of the instant polypeptide can be constructed by linking a gene or gene fragment encoding that polypeptide to plant promoter sequences. Alternatively, a chimeric gene designed to express antisense RNA for all or part of the instant nucleic acid fragment can be constructed by linking the gene or gene fragment in reverse orientation to plant promoter sequences. Either the co-suppression or antisense chimeric genes could be introduced into plants via transformation wherein expression of the corresponding endogenous genes are reduced or eliminated.  
       [0069] Molecular genetic solutions to the generation of plants with altered gene expression have a decided advantage over more traditional plant breeding approaches. Changes in plant phenotypes can be produced by specifically inhibiting expression of one or more genes by antisense inhibition or cosuppression (U.S. Pat. Nos. 5,190,931, 5,107,065 and 5,283,323). An antisense or cosuppression construct would act as a dominant negative regulator of gene activity. While conventional mutations can yield negative regulation of gene activity these effects are most likely recessive. The dominant negative regulation available with a transgenic approach may be advantageous from a breeding perspective. In addition, the ability to restrict the expression of specific phenotype to the reproductive tissues of the plant by the use of tissue specific promoters may confer agronomic advantages relative to conventional mutations which may have an effect in all tissues in which a mutant gene is ordinarily expressed.  
       [0070] The person skilled in the art will know that special considerations are associated with the use of antisense or cosuppression technologies in order to reduce expression of particular genes. For example, the proper level of expression of sense or antisense genes may require the use of different chimeric genes utilizing different regulatory elements known to the skilled artisan. Once transgenic plants are obtained by one of the methods described above, it will be necessary to screen individual transgenics for those that most effectively display the desired phenotype. Accordingly, the skilled artisan will develop methods for screening large numbers of transformants. The nature of these screens will generally be chosen on practical grounds, and is not an inherent part of the invention. For example, one can screen by looking for changes in gene expression by using antibodies specific for the protein encoded by the gene being suppressed, or one could establish assays that specifically measure enzyme activity. A preferred method will be one which allows large numbers of samples to be processed rapidly, since it will be expected that a large number of transformants will be negative for the desired phenotype.  
       [0071] The instant polypeptides (or portions thereof) may be produced in heterologous host cells, particularly in the cells of microbial hosts, and can be used to prepare antibodies to the these proteins by methods well known to those skilled in the art. The antibodies are useful for detecting the polypeptides of the instant invention in situ in cells or in vitro in cell extracts. Preferred heterologous host cells for production of the instant polypeptides are microbial hosts. Microbial expression systems and expression vectors containing regulatory sequences that direct high level expression of foreign proteins are well known to those skilled in the art. Any of these could be used to construct a chimeric gene for production of the instant polypeptides. This chimeric gene could then be introduced into appropriate microorganisms via transformation to provide high level expression of the encoded aminoacyl-tRNA synthetase. An example of a vector for high level expression of the instant polypeptides in a bacterial host is provided (Example 7).  
       [0072] Additionally, the instant polypeptides can be used as a targets to facilitate design and/or identification of inhibitors of those enzymes that may be useful as herbicides. This is desirable because the polypeptides described herein catalyze various steps in aminoacyl-tRNA biosynthesis. Accordingly, inhibition of the activity of one or more of the enzymes described herein could lead to inhibition of plant growth. Thus, the instant polypeptides could be appropriate for new herbicide discovery and design.  
       [0073] All or a substantial portion of the nucleic acid fragments of the instant invention may also be used as probes for genetically and physically mapping the genes that they are a part of, and as markers for traits linked to those genes. Such information may be useful in plant breeding in order to develop lines with desired phenotypes. For example, the instant nucleic acid fragments may be used as restriction fragment length polymorphism (RFLP) markers. Southern blots (Maniatis) of restriction-digested plant genomic DNA may be probed with the nucleic acid fragments of the instant invention. The resulting banding patterns may then be subjected to genetic analyses using computer programs such as MapMaker (Lander et al. (1987)  Genomics  1:174-181) in order to construct a genetic map. In addition, the nucleic acid fragments of the instant invention may be used to probe Southern blots containing restriction endonuclease-treated genomic DNAs of a set of individuals representing parent and progeny of a defined genetic cross. Segregation of the DNA polymorphisms is noted and used to calculate the position of the instant nucleic acid sequence in the genetic map previously obtained using this population (Botstein et al. (1980)  Am. J. Hum. Genet.  32:314-331).  
       [0074] The production and use of plant gene-derived probes for use in genetic mapping is described in Bernatzky and Tanksley (1986)  Plant Mol. Biol. Reporter  4:37-41. Numerous publications describe genetic mapping of specific cDNA clones using the methodology outlined above or variations thereof. For example, F2 intercross populations, backcross populations, randomly mated populations, near isogenic lines, and other sets of individuals may be used for mapping. Such methodologies are well known to those skilled in the art.  
       [0075] Nucleic acid probes derived from the instant nucleic acid sequences may also be used for physical mapping (i.e., placement of sequences on physical maps; see Hoheisel et al. In:  Nonmammalian Genomic Analysis: A Practical Guide,  Academic press 1996, pp. 319-346, and references cited therein).  
       [0076] In another embodiment, nucleic acid probes derived from the instant nucleic acid sequences may be used in direct fluorescence in situ hybridization (FISH) mapping (Trask (1991)  Trends Genet.  7:149-154). Although current methods of FISH mapping favor use of large clones (several to several hundred KB; see Laan et al. (1995)  Genome Res.  5:13-20), improvements in sensitivity may allow performance of FISH mapping using shorter probes.  
       [0077] A variety of nucleic acid amplification-based methods of genetic and physical mapping may be carried out using the instant nucleic acid sequences. Examples include allele-specific amplification (Kazazian (1989)  J. Lab. Clin. Med.  11:95-96), polymorphism of PCR-amplified fragments (CAPS; Sheffield et al. (1993)  Genomics  16:325-332), allele-specific ligation (Landegren et al. (1988)  Science  241:1077-1080), nucleotide extension reactions (Sokolov (1990)  Nucleic Acid Res.  18:3671), Radiation Hybrid Mapping (Walter et al. (1997)  Nat. Genet.  7:22-28) and Happy Mapping (Dear and Cook (1989)  Nucleic Acid Res.  17:6795-6807). For these methods, the sequence of a nucleic acid fragment is used to design and produce primer pairs for use in the amplification reaction or in primer extension reactions. The design of such primers is well known to those skilled in the art. In methods employing PCR-based genetic mapping, it may be necessary to identify DNA sequence differences between the parents of the mapping cross in the region corresponding to the instant nucleic acid sequence. This, however, is generally not necessary for mapping methods.  
       [0078] Loss of function mutant phenotypes may be identified for the instant cDNA clones either by targeted gene disruption protocols or by identifying specific mutants for these genes contained in a maize population carrying mutations in all possible genes (Ballinger and Benzer (1989)  Proc. Natl. Acad. Sci USA  86:9402-9406; Koes et al. (1995)  Proc. Natl. Acad. Sci USA  92:8149-8153; Bensen et al. (1995)  Plant Cell  7:75-84). The latter approach may be accomplished in two ways. First, short segments of the instant nucleic acid fragments may be used in polymerase chain reaction protocols in conjunction with a mutation tag sequence primer on DNAs prepared from a population of plants in which Mutator transposons or some other mutation-causing DNA element has been introduced (see Bensen, supra). The amplification of a specific DNA fragment with these primers indicates the insertion of the mutation tag element in or near the plant gene encoding the instant polypeptides. Alternatively, the instant nucleic acid fragment may be used as a hybridization probe against PCR amplification products generated from the mutation population using the mutation tag sequence primer in conjunction with an arbitrary genomic site primer, such as that for a restriction enzyme site-anchored synthetic adaptor. With either method, a plant containing a mutation in the endogenous gene encoding the instant polypeptides can be identified and obtained. This mutant plant can then be used to determine or confirm the natural function of the instant polypeptides disclosed herein. 
     
    
    
     EXAMPLES  
     [0079] The present invention is further defined in the following Examples, in which all parts and percentages are by weight and degrees are Celsius, unless otherwise stated. It should be understood that these Examples, while indicating preferred embodiments of the invention, are given by way of illustration only. From the above discussion and these Examples, one skilled in the art can ascertain the essential characteristics of this invention, and without departing from the spirit and scope thereof, can make various changes and modifications of the invention to adapt it to various usages and conditions.  
     Example 1  
     Composition of cDNA Libraries, Isolation and Sequencing of cDNA Clones  
     [0080] cDNA libraries representing mRNAs from various corn, rice, soybean and wheat tissues were prepared. The characteristics of the libraries are described below.  
               TABLE 2                          cDNA Libraries from Corn, Rice, Soybean and Wheat                         Library   Tissue   Clone               cr1n   Corn Root From 7 Day Old Seedlings*   cr1n.pk0140.b10       p0017   Corn Ear Shoot, Prophase I (2.8-4.8 cm)   p0017.cespf50r       res1c   Rice Etiolated Seedling   res1c.pk005.j13       r1r6   Rice Leaf 15 Days After Germination, 6 Hours After   r1r6.pk0084.e1           Infection of  Magaporthe grisea  Strain 4360-R-62           (AVR2-YAMO); Resistant               r1r6.pk0023.e9       s12   Soybean Two-Week-Old Developing Seedlings Treated   s12.pk123.g22           With 2.5 ppm chlorimuron       srr1c   Soybean 8-Day-Old Root   srr1c.pk003.k12       w1m4   Wheat Seedlings 4 Hours After Inoculation With  Erysiphe     w1m4.pk008.d5             graminis  f. sp tritici       w1m96   Wheat Seedlings 96 Hours After Inoculation With   w1m96.pk060.l13             Erysiphe               graminis  f. sp tritici       wr1   Wheat Root From 7 Day Old Seedling   wr1.pk181.e1                          
 
     [0081] cDNA libraries may be prepared by any one of many methods available. For example, the cDNAs may be introduced into plasmid vectors by first preparing the cDNA libraries in Uni-ZAP™ XR vectors according to the manufacturer&#39;s protocol (Stratagene Cloning Systems, La Jolla, Calif.). The Uni-ZAP™ XR libraries are converted into plasmid libraries according to the protocol provided by Stratagene. Upon conversion, cDNA inserts will be contained in the plasmid vector pBluescript. In addition, the cDNAs may be introduced directly into precut Bluescript II SK(+) vectors (Stratagene) using T4 DNA ligase (New England Biolabs), followed by transfection into DH10B cells according to the manufacturer&#39;s protocol (GIBCO BRL Products). Once the cDNA inserts are in plasmid vectors, plasmid DNAs are prepared from randomly picked bacterial colonies containing recombinant pBluescript plasmids, or the insert cDNA sequences are amplified via polymerase chain reaction using primers specific for vector sequences flanking the inserted cDNA sequences. Amplified insert DNAs or plasmid DNAs are sequenced in dye-primer sequencing reactions to generate partial cDNA sequences (expressed sequence tags or “ESTs”; see Adams et al., (1991)  Science  252:1651-1656). The resulting ESTs are analyzed using a Perkin Elmer Model 377 fluorescent sequencer.  
     Example 2  
     Identification of cDNA Clones  
     [0082] cDNA clones encoding aminoacyl-tRNA synthetases were identified by conducting BLAST (Basic Local Alignment Search Tool; Altschul et al. (1993)  J. Mol. Biol.  215:403-410; see also wwwv.nebi.nlm.nih.gov/BLAST/) searches for similarity to sequences contained in the BLAST “nr” database (comprising all non-redundant GenBank CDS translations, sequences derived from the 3-dimensional structure Brookhaven Protein Data Bank, the last major release of the SWISS-PROT protein sequence database, EMBL, and DDBJ databases). The cDNA sequences obtained in Example 1 were analyzed for similarity to all publicly available DNA sequences contained in the “nr” database using the BLASTN algorithm provided by the National Center for Biotechnology Information (NCBI). The DNA sequences were translated in all reading frames and compared for similarity to all publicly available protein sequences contained in the “nr” database using the BLASTX algorithm (Gish and States (1993)  Nat. Genet.  3:266-272) provided by the NCBI. For convenience, the P-value (probability) of observing a match of a cDNA sequence to a sequence contained in the searched databases merely by chance as calculated by BLAST are reported herein as “pLog” values, which represent the negative of the logarithm of the reported P-value. Accordingly, the greater the pLog value, the greater the likelihood that the cDNA sequence and the BLAST “hit” represent homologous proteins.  
     Example 3  
     Characterization of cDNA Clones Encoding Seryl-tRNA Synthetase  
     [0083] The BLASTX search using the EST sequences from clones listed in Table 3 revealed similarity of the polypeptides encoded by the cDNAs to seryl-tRNA synthetase from  Zea mays  (NCBI Identifier No. gi 3319776) and  Arabidopsis thaliana  (NCBI Identifier No. gi 2501056). Shown in Table 3 are the BLAST results for individual ESTs (“EST”), the sequences of the entire cDNA inserts comprising the indicated cDNA clones (“FIS”), contigs assembled from two or more ESTs (“Contig”), contigs assembled from an FIS and one or more ESTs (“Contig*”), or sequences encoding the entire protein derived from an FIS, a contig, or an FIS and PCR (“CGS”):  
               TABLE 3                          BLAST Results for Sequences Encoding Polypeptides Homologous       to  Zea mays  and  Arabidopsis thaliana  Sery1-tRNA Synthetase                                 Clone   Status   BLAST pLog Score                       r1r6.pk0023.e9   CGS   &gt;254.00 (gi 3319776)           s12.pk123.g22   FIS      51.00 (gi 2501056)           w1m4.pk0018.d5   CGS   &gt;254.00 (gi 3319776)                      
 
     [0084] The data in Table 4 represents a calculation of the percent identity of the amino acid sequences set forth in SEQ ID NOs:16, 18 and 20 and the  Zea mays  and  Arabidopsis thaliana  sequences.  
               TABLE 4                          Percent Identity of Amino Acid Sequences Deduced From the       Nucleotide Sequences of cDNA Clones Encoding Polypeptides       Homologous to  Zea mays  and  Arabidoesis thaliana         Seryl-tRNA Synthetase                             SEQ ID NO.   Percent Identity to                       16   75% (gi 3319776)           18   80% (gi 2501056)           20   80% (gi 3319776)                      
 
     [0085] Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal method of alignment (Higgins and Sharp (1989)  CABIOS.  5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. Sequence alignments and BLAST scores and probabilities indicate that the nucleic acid fragments comprising the instant cDNA clones encode a substantial portion of a seryl-tRNA synthetase. These sequences represent the first rice, soybean and wheat sequences encoding seryl-tRNA synthetase.  
     Example 4  
     Characterization of cDNA Clones Encoding Threonyl-tRNA Synthetase  
     [0086] The BLASTX search using the EST sequences from clones listed in Table 5 revealed similarity of the polypeptides encoded by the cDNAs to threonyl-tRNA synthetase from  Arabidopsis thaliana  (NCBI Identifier No. gi 3617770). Shown in Table 5 are the BLAST results for individual ESTs (“EST”), the sequences of the entire cDNA inserts comprising the indicated cDNA clones (“FIS”), contigs assembled from two or more ESTs (“Contig”), contigs assembled from an FIS and one or more ESTs (“Contig*”), or sequences encoding the entire protein derived from an FIS, a contig, or an FIS and PCR (“CGS”):  
               TABLE 5                          BLAST Results for Sequences Encoding Polypeptides Homologous       to  Arabidopis thaliana  Threonyl-t-RNA Synthetase                         Clone   Status   BLAST pLog Score to gi 3617770               p0017.cespf50r   FIS   &gt;254.00       res1c.pk005.j13   FIS   &gt;254.00       srr1c.pk003.k12   FIS   &gt;254.00       w1m96.pk060.l13   FIS   &gt;254.00                  
 
     [0087] The data in Table 6 represents a calculation of the percent identity of the amino acid sequences set forth in SEQ ID NOs:22, 24, 26 and 28 and the  Arabidopsis thaliana  sequence.  
               TABLE 6                          Percent Identity of Amino Acid Sequences Deduced From the Nucleotide       Sequences of cDNA Clones Encoding Polypeptides Homologous to\         Arabidopsis thaliana  Threonyl-tRNA Synthetase                             SEQ ID NO.   Percent Identity to                       22   66%           24   73%           26   76%           28   76%                      
 
     [0088] Sequence alignments and percent identity calculations were performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR Inc., Madison, Wis.). Multiple alignment of the sequences was performed using the Clustal method of alignment (Higgins and Sharp (1989)  CABIOS.  5:151-153) with the default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Default parameters for pairwise alignments using the Clustal method were KTUPLE 1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5. Sequence alignments and BLAST scores and probabilities indicate that the nucleic acid fragments comprising the instant cDNA clones encode a substantial portion of a threonyl-tRNA synthetase. These sequences represent the first corn, rice, soybean and wheatsequences encoding threonyl-tRNA synthetase.  
     Example 5  
     Expression of Chimeric Genes in Monocot Cells  
     [0089] A chimeric gene comprising a cDNA encoding the instant polypeptide in sense orientation with respect to the maize 27 kD zein promoter that is located 5′ to the cDNA fragment, and the 10 kD zein 3′ end that is located 3′ to the cDNA fragment, can be constructed. The cDNA fragment of this gene may be generated by polymerase chain reaction (PCR) of the cDNA clone using appropriate oligonucleotide primers. Cloning sites (NcoI or SmaI) can be incorporated into the oligonucleotides to provide proper orientation of the DNA fragment when inserted into the digested vector pML103 as described below. Amplification is then performed in a standard PCR. The amplified DNA is then digested with restriction enzymes NcoI and SmaI and fractionated on an agarose gel. The appropriate band can be isolated from the gel and combined with a 4.9 kb NcoI-SmaI fragment of the plasmid pML103. Plasmid pML103 has been deposited under the terms of the Budapest Treaty at ATCC (American Type Culture Collection, 10801 University Blvd., Manassas, Va. 20110-2209), and bears accession number ATCC 97366. The DNA segment from pML103 contains a 1.05 kb SalI-NcoI promoter fragment of the maize 27 kD zein gene and a 0.96 kb SmaI-SalI fragment from the 3′ end of the maize 10 kD zein gene in the vector pGem9Zf(+) (Promega). Vector and insert DNA can be ligated at 15° C. overnight, essentially as described (Maniatis). The ligated DNA may then be used to transform  E. coli  XL1-Blue (Epicurian Coli XL-1 Blue™; Stratagene). Bacterial transformants can be screened by restriction enzyme digestion of plasmid DNA and limited nucleotide sequence analysis using the dideoxy chain termination method (Sequenase™ DNA Sequencing Kit; U.S. Biochemical). The resulting plasmid construct would comprise a chimeric gene encoding, in the 5′ to 3′ direction, the maize 27 kD zein promoter, a cDNA fragment encoding the instant polypeptides, and the 10 kD zein 3′ region.  
     [0090] The chimeric gene described above can then be introduced into corn cells by the following procedure. Immature corn embryos can be dissected from developing caryopses derived from crosses of the inbred corn lines H99 and LH132. The embryos are isolated 10 to 11 days after pollination when they are 1.0 to 1.5 mm long. The embryos are then placed with the axis-side facing down and in contact with agarose-solidified N6 medium (Chu et al. (1975)  Sci. Sin. Peking  18:659-668). The embryos are kept in the dark at 27° C. Friable embryogenic callus consisting of undifferentiated masses of cells with somatic proembryoids and embryoids borne on suspensor structures proliferates from the scutellum of these immature embryos. The embryogenic callus isolated from the primary explant can be cultured on N6 medium and sub-cultured on this medium every 2 to 3 weeks.  
     [0091] The plasmid, p35S/Ac (obtained from Dr. Peter Eckes, Hoechst Ag, Frankfurt, Germany) may be used in transformation experiments in order to provide for a selectable marker. This plasmid contains the Pat gene (see European Patent Publication 0 242 236) which encodes phosphinothricin acetyl transferase (PAT). The enzyme PAT confers resistance to herbicidal glutamine synthetase inhibitors such as phosphinothricin. The pat gene in p35S/Ac is under the control of the 35S promoter from Cauliflower Mosaic Virus (Odell et al. (1985)  Nature  313:810-812) and the 3′ region of the nopaline synthase gene from the T-DNA of the Ti plasmid of  Agrobacterium tumefaciens.    
     [0092] The particle bombardment method (Klein et al. (1987)  Nature  327:70-73) may be used to transfer genes to the callus culture cells. According to this method, gold particles (1 μm in diameter) are coated with DNA using the following technique. Ten μg of plasmid DNAs are added to 50 μ  of a suspension of gold particles (60 mg per m ). Calcium chloride (50 μ  of a 2.5 M solution) and spermidine free base (20 μ  of a 1.0 M solution) are added to the particles. The suspension is vortexed during the addition of these solutions. After 10 minutes, the tubes are briefly centrifuged (5 sec at 15,000 rpm) and the supernatant removed. The particles are resuspended in 200 μ  of absolute ethanol, centrifuged again and the supernatant removed. The ethanol rinse is performed again and the particles resuspended in a final volume of 30 μ  of ethanol. An aliquot (5 μ  ) of the DNA-coated gold particles can be placed in the center of a Kapton™ flying disc (Bio-Rad Labs). The particles are then accelerated into the corn tissue with a Biolistic™ PDS-1000/He (Bio-Rad Instruments, Hercules Calif.), using a helium pressure of 1000 psi, a gap distance of 0.5 cm and a flying distance of 1.0 cm.  
     [0093] For bombardment, the embryogenic tissue is placed on filter paper over agarose-solidified N6 medium. The tissue is arranged as a thin lawn and covered a circular area of about 5 cm in diameter. The petri dish containing the tissue can be placed in the chamber of the PDS-1000/He approximately 8 cm from the stopping screen. The air in the chamber is then evacuated to a vacuum of 28 inches of Hg. The macrocarrier is accelerated with a helium shock wave using a rupture membrane that bursts when the He pressure in the shock tube reaches 1000 psi.  
     [0094] Seven days after bombardment the tissue can be transferred to N6 medium that contains gluphosinate (2 mg per liter) and lacks casein or proline. The tissue continues to grow slowly on this medium. After an additional 2 weeks the tissue can be transferred to fresh N6 medium containing gluphosinate. After 6 weeks, areas of about 1 cm in diameter of actively growing callus can be identified on some of the plates containing the glufosinate-supplemented medium. These calli may continue to grow when sub-cultured on the selective medium.  
     [0095] Plants can be regenerated from the transgenic callus by first transferring clusters of tissue to N6 medium supplemented with 0.2 mg per liter of 2,4-D. After two weeks the tissue can be transferred to regeneration medium (Fromm et al. (1990)  Bio/Technology  8:833-839).  
     Example 6  
     Expression of Chimeric Genes in Dicot Cells  
     [0096] A seed-specific expression cassette composed of the promoter and transcription terminator from the gene encoding the β subunit of the seed storage protein phaseolin from the bean  Phaseolus vulgaris  (Doyle et al. (1986)  J. Biol. Chem.  261:9228-9238) can be used for expression of the instant polypeptides in transformed soybean. The phaseolin cassette includes about 500 nucleotides upstream (5′) from the translation initiation codon and about 1650 nucleotides downstream (3′) from the translation stop codon of phaseolin. Between the 5′ and 3′ regions are the unique restriction endonuclease sites Nco I (which includes the ATG translation initiation codon), Sma I, Kpn I and Xba I. The entire cassette is flanked by Hind III sites.  
     [0097] The cDNA fragment of this gene may be generated by polymerase chain reaction (PCR) of the cDNA clone using appropriate oligonucleotide primers. Cloning sites can be incorporated into the oligonucleotides to provide proper orientation of the DNA fragment when inserted into the expression vector. Amplification is then performed as described above, and the isolated fragment is inserted into a pUC18 vector carrying the seed expression cassette.  
     [0098] Soybean embryos may then be transformed with the expression vector comprising sequences encoding the instant polypeptides. To induce somatic embryos, cotyledons, 3-5 mm in length dissected from surface sterilized, immature seeds of the soybean cultivar A2872, can be cultured in the light or dark at 26° C. on an appropriate agar medium for 6-10 weeks. Somatic embryos which produce secondary embryos are then excised and placed into a suitable liquid medium. After repeated selection for clusters of somatic embryos which multiplied as early, globular staged embryos, the suspensions are maintained as described below.  
     [0099] Soybean embryogenic suspension cultures can maintained in 35 m  liquid media on a rotary shaker, 150 rpm, at 26° C. with florescent lights on a 16:8 hour day/night schedule. Cultures are subcultured every two weeks by inoculating approximately 35 mg of tissue into 35 m  of liquid medium.  
     [0100] Soybean embryogenic suspension cultures may then be transformed by the method of particle gun bombardment (Klein et al. (1987)  Nature  (London) 327:70-73, U.S. Pat. No. 4,945,050). A DuPont Biolistic™ PDS 1000/HE instrument (helium retrofit) can be used for these transformations.  
     [0101] A selectable marker gene which can be used to facilitate soybean transformation is a chimeric gene composed of the 35S promoter from Cauliflower Mosaic Virus (Odell et al. (1985)  Nature  313:810-812), the hygromycin phosphotransferase gene from plasmid pJR225 (from  E. coli;  Gritz et al.(1983)  Gene  25:179-188) and the 3′ region of the nopaline synthase gene from the T-DNA of the Ti plasmid of  Agrobacterium tumefaciens.  The seed expression cassette comprising the phaseolin 5′ region, the fragment encoding the instant polypeptides and the phaseolin 3′ region can be isolated as a restriction fragment. This fragment can then be inserted into a unique restriction site of the vector carrying the marker gene.  
     [0102] To 50 μL of a 60 mg/m  1 μm gold particle suspension is added (in order): 5 μ  DNA (1 μg/μ ), 20 μl spermidine (0.1 M), and 50 μ  CaCl 2  (2.5 M). The particle preparation is then agitated for three minutes, spun in a microfuge for 10 seconds and the supernatant removed. The DNA-coated particles are then washed once in 400 μ  70% ethanol and resuspended in 40 μ  of anhydrous ethanol. The DNA/particle suspension can be sonicated three times for one second each. Five μ  of the DNA-coated gold particles are then loaded on each macro carrier disk.  
     [0103] Approximately 300-400 mg of a two-week-old suspension culture is placed in an empty 60×15 mm petri dish and the residual liquid removed from the tissue with a pipette. For each transformation experiment, approximately 5-10 plates of tissue are normally bombarded. Membrane rupture pressure is set at 1100 psi and the chamber is evacuated to a vacuum of 28 inches mercury. The tissue is placed approximately 3.5 inches away from the retaining screen and bombarded three times. Following bombardment, the tissue can be divided in half and placed back into liquid and cultured as described above.  
     [0104] Five to seven days post bombardment, the liquid media may be exchanged with fresh media, and eleven to twelve days post bombardment with fresh media containing 50 mg/m  hygromycin. This selective media can be refreshed weekly. Seven to eight weeks post bombardment, green, transformed tissue may be observed growing from untransformed, necrotic embryogenic clusters. Isolated green tissue is removed and inoculated into individual flasks to generate new, clonally propagated, transformed embryogenic suspension cultures. Each new line may be treated as an independent transformation event. These suspensions can then be subcultured and maintained as clusters of immature embryos or regenerated into whole plants by maturation and germination of individual somatic embryos.  
     Example 7  
     Expression of Chimeric Genes in Microbial Cells  
     [0105] The cDNAs encoding the instant polypeptides can be inserted into the T7  E. coli  expression vector pBT430. This vector is a derivative of pET-3a (Rosenberg et al. (1987)  Gene  56:125-135) which employs the bacteriophage T7 RNA polymerase/T7 promoter system. Plasmid pBT430 was constructed by first destroying the EcoR I and Hind III sites in pET-3a at their original positions. An oligonucleotide adaptor containing EcoR I and Hind III sites was inserted at the BamH I site of pET-3a. This created pET-3aM with additional unique cloning sites for insertion of genes into the expression vector. Then, the Nde I site at the position of translation initiation was converted to an Nco I site using oligonucleotide-directed mutagenesis. The DNA sequence of pET-3aM in this region, 5′-CATATGG, was converted to 5′-CCCATGG in pBT430.  
     [0106] Plasmid DNA containing a cDNA may be appropriately digested to release a nucleic acid fragment encoding the protein. This fragment may then be purified on a 1% NuSieve GTG™ low melting agarose gel (FMC). Buffer and agarose contain 10 μg/ml ethidium bromide for visualization of the DNA fragment. The fragment can then be purified from the agarose gel by digestion with GELase™ (Epicentre Technologies) according to the manufacturer&#39;s instructions, ethanol precipitated, dried and resuspended in 20 μ  of water. Appropriate oligonucleotide adapters may be ligated to the fragment using T4 DNA ligase (New England Biolabs, Beverly, Mass.). The fragment containing the ligated adapters can be purified from the excess adapters using low melting agarose as described above. The vector pBT430 is digested, dephosphorylated with alkaline phosphatase (NEB) and deproteinized with phenol/chloroform as described above. The prepared vector pBT430 and fragment can then be ligated at 16° C. for 15 hours followed by transformation into DH5 electrocompetent cells (GIBCO BRL). Transformants can be selected on agar plates containing LB media and 100 μg/m  ampicillin. Transformants containing the gene encoding the instant polypeptides are then screened for the correct orientation with respect to the T7 promoter by restriction enzyme analysis.  
     [0107] For high level expression, a plasmid clone with the cDNA insert in the correct orientation relative to the T7 promoter can be transformed into  E. coli  strain BL21(DE3) (Studier et al. (1986)  J. Mol. Biol.  189:113-130). Cultures are grown in LB medium containing ampicillin (100 mg/L) at 25° C. At an optical density at 600 nm of approximately 1, IPTG (isopropylthio-β-galactoside, the inducer) can be added to a final concentration of 0.4 mM and incubation can be continued for 3 h at 25°. Cells are then harvested by centrifugation and re-suspended in 50 μ  of 50 mM Tris-HCl at pH 8.0 containing 0.1 mM DTT and 0.2 mM phenyl methylsulfonyl fluoride. A small amount of 1 mm glass beads can be added and the mixture sonicated 3 times for about 5 seconds each time with a microprobe sonicator. The mixture is centrifuged and the protein concentration of the supernatant determined. One μg of protein from the soluble fraction of the culture can be separated by SDS-polyacrylamide gel electrophoresis. Gels can be observed for protein bands migrating at the expected molecular weight.  
     Example 8  
     Evaluating Compounds for Their Ability to Inhibit the Activity of Aminoacyl-tRNA Synthetases  
     [0108] The polypeptides described herein may be produced using any number of methods known to those skilled in the art. Such methods include, but are not limited to, expression in bacteria as described in Example 7, or expression in eukaryotic cell culture, in planta, and using viral expression systems in suitably infected organisms or cell lines. The instant polypeptides may be expressed either as mature forms of the proteins as observed in vivo or as fusion proteins by covalent attachment to a variety of enzymes, proteins or affinity tags. Common fusion protein partners include glutathione S-transferase (“GST”), thioredoxin (“Trx”), maltose binding protein, and C- and/or N-terminal hexahistidine polypeptide (“(His) 6 ”). The fusion proteins may be engineered with a protease recognition site at the fusion point so that fusion partners can be separated by protease digestion to yield intact mature enzyme. Examples of such proteases include thrombin, enterokinase and factor Xa. However, any protease can be used which specifically cleaves the peptide connecting the fusion protein and the enzyme.  
     [0109] Purification of the instant polypeptides, if desired, may utilize any number of separation technologies familiar to those skilled in the art of protein purification. Examples of such methods include, but are not limited to, homogenization, filtration, centrifugation, heat denaturation, ammonium sulfate precipitation, desalting, pH precipitation, ion exchange chromatography, hydrophobic interaction chromatography and affinity chromatography, wherein the affinity ligand represents a substrate, substrate analog or inhibitor. When the instant polypeptides are expressed as fusion proteins, the purification protocol may include the use of an affinity resin which is specific for the fusion protein tag attached to the expressed enzyme or an affinity resin containing ligands which are specific for the enzyme. For example, the instant polypeptides may be expressed as a fusion protein coupled to the C-terminus of thioredoxin. In addition, a (His) 6  peptide may be engineered into the N-terminus of the fused thioredoxin moiety to afford additional opportunities for affinity purification. Other suitable affinity resins could be synthesized by linking the appropriate ligands to any suitable resin such as Sepharose-4B. In an alternate embodiment, a thioredoxin fusion protein may be eluted using dithiothreitol; however, elution may be accomplished using other reagents which interact to displace the thioredoxin from the resin. These reagents include β-mercaptoethanol or other reduced thiol. The eluted fusion protein may be subjected to further purification by traditional means as stated above, if desired. Proteolytic cleavage of the thioredoxin fusion protein and the enzyme may be accomplished after the fusion protein is purified or while the protein is still bound to the ThioBond™ affinity resin or other resin.  
     [0110] Crude, partially purified or purified enzyme, either alone or as a fusion protein, may be utilized in assays for the evaluation of compounds for their ability to inhibit enzymatic activation of the instant polypeptides disclosed herein. Assays may be conducted under well known experimental conditions which permit optimal enzymatic activity. For example, assays for aminoacyl-tRNA synthetase activity are presented by (Lloyd et al., (1995)  Nucleic Acid Research  23(15):2882-2892).  
    
     
       
         1 
         
           
             28  
           
           
             1  
             562  
             DNA  
             Oryza sp.  
             
               unsure  
               (431)  
               n = a, c, g or t  
             
           
            1 

gtttaaactc tacccttccc tctctgctcg ccgccgccgc cgccgccgca cgccttcgaa     60 

gatgctcgac atcaacctct tccgcacgga gaagggcggc gacccggagc tgatccgcag    120 

gtcgcagcgc aaccgctccg cctccgtcga gctcgtcgac gaggtcatcg ccctcgacga    180 

ccagtggcgc cagaggcaat tcgagctcga caaaatccgg caggagctca acaaaaccag    240 

caaggaaatc ggcaagctca aggctaaaaa ggaggacgcg tcggctctga ttcagagcac    300 

ggaagagatt aagaagaggt tggctgccaa ggagacggag gtgcaggaag ccaagggcac    360 

gctcgatgcc aagctcgtga cgattggaaa cattgtgcat gaatccgtcc ctgtcagcga    420 

tgacgaggct ncaatttaat tgtacggaca tggggaagag aggctggagg aatttgaaga    480 

nacgtggatt tngaaatntt acatagagct tgagangtnt ntgatcangt gaanggtaca    540 

ttaatganna gggccactga ct                                             562 

 
           
             2  
             125  
             PRT  
             Oryza sp.  
             
               UNSURE  
               (124)  
               Xaa = any amino acid  
             
           
            2 

Met Leu Asp Ile Asn Leu Phe Arg Thr Glu Lys Gly Gly Asp Pro Glu 
  1               5                  10                  15 

Leu Ile Arg Arg Ser Gln Arg Asn Arg Ser Ala Ser Val Glu Leu Val 
             20                  25                  30 

Asp Glu Val Ile Ala Leu Asp Asp Gln Trp Arg Gln Arg Gln Phe Glu 
         35                  40                  45 

Leu Asp Lys Ile Arg Gln Glu Leu Asn Lys Thr Ser Lys Glu Ile Gly 
     50                  55                  60 

Lys Leu Lys Ala Lys Lys Glu Asp Ala Ser Ala Leu Ile Gln Ser Thr 
 65                  70                  75                  80 

Glu Glu Ile Lys Lys Arg Leu Ala Ala Lys Glu Thr Glu Val Gln Glu 
                 85                  90                  95 

Ala Lys Gly Thr Leu Asp Ala Lys Leu Val Thr Ile Gly Asn Ile Val 
            100                 105                 110 

His Glu Ser Val Pro Val Ser Asp Asp Glu Ala Xaa Ile 
        115                 120                 125 

 
           
             3  
             489  
             DNA  
             Glycine max  
           
            3 

atgatgctgc agcaaagaag tatgatctag aagcatggtt tccagcctct caagcttaca     60 

gagagctagt gtcctgttca aactgtacag actatcaggc cagaagatta gaaattcgat    120 

atggtcagaa aaagagcaat gagcaaatga agcaatatgt tcacttgttg aactctactc    180 

taacggctac tgagaggacc atttgctgca tactagagaa caaccagaag gaagatgggg    240 

tagagatacc agaagccctc aggccattca tgggtggaaa gactttccta cctttcaaga    300 

accaaccatc taatgaagcc aaagggaaga aatcgaaggc ctaattgcat tttaagtcag    360 

tgatatttat gaagagtttg ttcagagttg taggttatga tgaccccgat attatccacc    420 

acacactgtt tccctttctc actaaatagt aaaatgattt ttaggagcac gcaccatttt    480 

tgggtcaaa                                                            489  
           
             4  
             102  
             PRT  
             Glycine max  
           
            4 

Asp Ala Ala Ala Lys Lys Tyr Asp Leu Glu Ala Trp Phe Pro Ala Ser 
  1               5                  10                  15 

Gln Ala Tyr Arg Glu Leu Val Ser Cys Ser Asn Cys Thr Asp Tyr Gln 
             20                  25                  30 

Ala Arg Arg Leu Glu Ile Arg Tyr Gly Gln Lys Lys Ser Asn Glu Gln 
         35                  40                  45 

Met Lys Gln Tyr Val His Leu Leu Asn Ser Thr Leu Thr Ala Thr Glu 
     50                  55                  60 

Arg Thr Ile Cys Cys Ile Leu Glu Asn Asn Gln Lys Glu Asp Gly Val 
 65                  70                  75                  80 

Glu Ile Pro Glu Ala Leu Arg Pro Phe Met Gly Gly Lys Thr Phe Leu 
                 85                  90                  95 

Pro Phe Lys Asn Gln Pro 
            100 

 
           
             5  
             641  
             DNA  
             Triticum sp.  
             
               unsure  
               (354)  
               n = a, c, g or t  
             
           
            5 

ccgtcatcca cctctcgtca gaagaaccct ccctctccgc cctccgccgc cgccgccgcc     60 

gccgccaatc gcagcgcagc cgcctcgccc ccgtcgagct cgtccgcgga gaagttggcc    120 

tagccgcggt cgcacgagaa tatgctcgac atcaacctct tccgcaagga gaagggcggc    180 

gaccctgagc tcgtccgcca gtcgcagcgc agccgcttcg cccccgtcga gctcgtcgac    240 

gaggtcatcg tcctcgacga ggcgtggcgc cagaggcagt tcgagctcga caagatccgg    300 

caggagctca acaaaaccag caaggagatc ggcaagctca aggccaaaaa gcangatgcg    360 

acggagctga tacagagcac ggagggagat taagaagagg ctggccgcca aggagacgga    420 

cgtncaggag gcaagacacc tcgatgcaag ctagttacat cggcaactcg tgcatgatct    480 

gnccataaca acgacgagca aacatctttg ncggcatggg caanaaanat ggggagaatn    540 

aagatcagtg gtnctnnaaa tgctgnanct ancttgaaag gnctagttct gggaaaggtn    600 

ntttaaggga nggttccnaa cangttaaaa tttggcatat n                        641 

 
           
             6  
             81  
             PRT  
             Triticum sp.  
             
               UNSURE  
               (71)  
               Xaa = any amino acid  
             
           
            6 

Met Leu Asp Ile Asn Leu Phe Arg Lys Glu Lys Gly Gly Asp Pro Glu 
  1               5                  10                  15 

Leu Val Arg Gln Ser Gln Arg Ser Arg Phe Ala Pro Val Glu Leu Val 
             20                  25                  30 

Asp Glu Val Ile Val Leu Asp Glu Ala Trp Arg Gln Arg Gln Phe Glu 
         35                  40                  45 

Leu Asp Lys Ile Arg Gln Glu Leu Asn Lys Thr Ser Lys Glu Ile Gly 
     50                  55                  60 

Lys Leu Lys Ala Lys Lys Xaa Asp Ala Thr Glu Leu Ile Gln Ser Thr 
 65                  70                  75                  80 

Glu 

 
           
             7  
             536  
             DNA  
             Zea mays  
             
               unsure  
               (333)  
               n = a, c, g or t  
             
           
            7 

gaaacttgga acaaagctga gcaacaattg acagaagctt taaatgagtt tgggaagcca     60 

tggaagatta atgaagggga tggtgctttc tacggcccaa aaattgatat tggtgtgttt    120 

gatgccctta agaggaaatt tcagtgtgca accctacagc tggattttca gctgcccatt    180 

cggttcaagc tggcttattc tgctgaggat gaagccaaaa ttgaaaggcc tgtgatgata    240 

cacagggcaa tcctaggttc ggttgaaagg atgcttgcca ttcttttggg agcattacaa    300 

tggtaaatgg ccttatggct aaccctcgcc agncattgtt tgctcggtac ttctggtcag    360 

tggatatgcc aaacaantcc tgccactcca catgaactgg tttcatgttg atattgaccc    420 

aatgacanga caatacaaaa gaagnacngg aactcaactg gccaatcaac tacatcctgt    480 

cgtagtgcac aanaagcnag acgggaatat atccttaggg aanagaaatc nactgt        536 

 
           
             8  
             95  
             PRT  
             Zea mays  
           
            8 

Glu Thr Trp Asn Lys Ala Glu Gln Gln Leu Thr Glu Ala Leu Asn Glu 
  1               5                  10                  15 

Phe Gly Lys Pro Trp Lys Ile Asn Glu Gly Asp Gly Ala Phe Tyr Gly 
             20                  25                  30 

Pro Lys Ile Asp Ile Gly Val Phe Asp Ala Leu Lys Arg Lys Phe Gln 
         35                  40                  45 

Cys Ala Thr Leu Gln Leu Asp Phe Gln Leu Pro Ile Arg Phe Lys Leu 
     50                  55                  60 

Ala Tyr Ser Ala Glu Asp Glu Ala Lys Ile Glu Arg Pro Val Met Ile 
 65                  70                  75                  80 

His Arg Ala Ile Leu Gly Ser Val Glu Arg Met Leu Ala Ile Leu 
                 85                  90                  95 

 
           
             9  
             425  
             DNA  
             Oryza sp.  
             
               unsure  
               (249)  
               n = a, c, g or t  
             
           
            9 

gtttaaacga ttcatagagc tatccttggg tctttggaac gattttttgg tgtcctcatt     60 

gaacactatg ctggtgattt tccactttgg cttgcaccaa tccaagctcg tattctacct    120 

gtgacagaca atgagctgca atactgtaac gaggtggctt cagaactgaa atcaaaaggc    180 

attccgagct gaggtatgtc atggcgagcg tctaccaaag ctaatacgga atgcccgaaa    240 

cgaagaaant gccgctcatg ggcggccttg gggcctaaag aantcnaagc aaggaccctc    300 

cactacaggc caagcatagt ggggaattgg gactatgcct gtgggatgat tccttcgcag    360 

aaccaacttg ctattggcga acaacctcct caactaatga acatttaaat atgctaaaga    420 

cagtt                                                                425 

 
           
             10  
             58  
             PRT  
             Oryza sp.  
           
            10 

Ile His Arg Ala Ile Leu Gly Ser Leu Glu Arg Phe Phe Gly Val Leu 
  1               5                  10                  15 

Ile Glu His Tyr Ala Gly Asp Phe Pro Leu Trp Leu Ala Pro Ile Gln 
             20                  25                  30 

Ala Arg Ile Leu Pro Val Thr Asp Asn Glu Leu Gln Tyr Cys Asn Glu 
         35                  40                  45 

Val Ala Ser Glu Leu Lys Ser Lys Gly Ile 
     50                  55 

 
           
             11  
             508  
             DNA  
             Glycine max  
             
               unsure  
               (467)  
               n = a, c, g or t  
             
           
            11 

acactatgct atgctatgct ctctaatccg tttccgccgt tacgctcctt cctaccgcac     60 

tctccactct ctctttccga cgattaaccg tttctcctcc tccgtctcct ccgcctccgc    120 

cgccatggtt gctcacgcga aggacgaggc gtacctcagc gcgacgattc cgaaacgcat    180 

ccgtctcttc gagaccatcc tggcggagca gcacactcag cgcctctcgc tctccccgga    240 

tcctatcaag gttactctcc ccgacggcag cgtcaaggag gcgaagaagt ggcatacgac    300 

gccgcttgat gttgcgcgtg aaatctcgaa gaatttggcc aacagcgcgc tcatcgcgaa    360 

ggtcaatggc gtgctctggg acatgactcg ccctctcgag gacgattgcc aagctccaga    420 

tcttcaagtt cgacgacgac gaaggccgcg acaccttctg ggactcnagc gcccacattc    480 

tcgggcaagt cacttgagac ggatatgg                                       508 

 
           
             12  
             80  
             PRT  
             Glycine max  
           
            12 

Ile Pro Lys Arg Ile Arg Leu Phe Glu Thr Ile Leu Ala Glu Gln His 
  1               5                  10                  15 

Thr Gln Arg Leu Ser Leu Ser Pro Asp Pro Ile Lys Val Thr Leu Pro 
             20                  25                  30 

Asp Gly Ser Val Lys Glu Ala Lys Lys Trp His Thr Thr Pro Leu Asp 
         35                  40                  45 

Val Ala Arg Glu Ile Ser Lys Asn Leu Ala Asn Ser Ala Leu Ile Ala 
     50                  55                  60 

Lys Val Asn Gly Val Leu Trp Asp Met Thr Arg Pro Leu Glu Asp Asp 
 65                  70                  75                  80 

 
           
             13  
             487  
             DNA  
             Triticum sp.  
             
               unsure  
               (467)  
               n = a, c, g or t  
             
           
            13 

gttattaggg cagtcccaga aactcttctt tttccatcca cttagcccag gtagctgctt     60 

cttccttcca aatggcgcta taatatataa caaattgatg gattttttgc gcaaggagta    120 

tagagagaga ggctaccaag aggttctgag tccaaatatt tacaacatgc aactttggga    180 

aacctctgga catgctgcaa actacaagga caacatgttt gtttttgaga tcgagaaaca    240 

agaatttggc cttaagccaa tgaattgtcc tggccattgc ctaatgtttg gacacgaggt    300 

tcgatcgtat agagagttgc ctctccgcat ggctgatttt gggagttctg cacagaaatg    360 

aacttagtgg gtgcacttac aggtttgaca cgtgtcagaa gatccaacag gacgatgccc    420 

atattttttg cacggagagc aaatcaagga tgaagttcgg gcttgcntgg gagtcaatga    480 

tatgtta                                                              487 

 
           
             14  
             146  
             PRT  
             Triticum sp.  
           
            14 

Leu Gly Gln Ser Gln Lys Leu Phe Phe Phe His Pro Leu Ser Pro Gly 
  1               5                  10                  15 

Ser Cys Phe Phe Leu Pro Asn Gly Ala Ile Ile Tyr Asn Lys Leu Met 
             20                  25                  30 

Asp Phe Leu Arg Lys Glu Tyr Arg Glu Arg Gly Tyr Gln Glu Val Leu 
         35                  40                  45 

Ser Pro Asn Ile Tyr Asn Met Gln Leu Trp Glu Thr Ser Gly His Ala 
     50                  55                  60 

Ala Asn Tyr Lys Asp Asn Met Phe Val Phe Glu Ile Glu Lys Gln Glu 
 65                  70                  75                  80 

Phe Gly Leu Lys Pro Met Asn Cys Pro Gly His Cys Leu Met Phe Gly 
                 85                  90                  95 

His Glu Val Arg Ser Tyr Arg Glu Leu Pro Leu Arg Met Ala Asp Phe 
            100                 105                 110 

Gly Val Leu His Arg Asn Glu Leu Ser Gly Cys Thr Tyr Arg Phe Asp 
        115                 120                 125 

Thr Cys Gln Lys Ile Gln Gln Asp Asp Ala His Ile Phe Cys Thr Glu 
    130                 135                 140 

Ser Lys 
145 

 
           
             15  
             1753  
             DNA  
             Oryza sativa  
           
            15 

gcacgaggtt taaactctac ccttccctct ctgctcgccg ccgccgccgc cgccgcacgc     60 

cttcgaagat gctcgacatc aacctcttcc gcacggagaa gggcggcgac ccggagctga    120 

tccgcaggtc gcagcgcaac cgctccgcct ccgtcgagct cgtcgacgag gtcatcgccc    180 

tcgacgacca gtggcgccag aggcaattcg agctcgacaa aatccggcag gagctcaaca    240 

aaaccagcaa ggaaatcggc aagctcaagg ctaaaaagga ggacgcgtcg gctctgattc    300 

agagcacgga agagattaag aagaggttgg ctgccaagga gacggaggtg caggaggcca    360 

agggcacgct cgatgccaag ctcgtgacga ttggaaacat tgtgcatgaa tccgtccctg    420 

tcagcgatga cgaggctaac aatttaattg tacggacatg gggagagagg aggctggagg    480 

gtaatttgaa gaatcacgtg gatctttgta agatgcttga catagtagct ttggagaaag    540 

gtgctgatgt agcaggtgga aggggttact atttaaagga tgaaggtgtc ctactgaact    600 

tggcattgat aaattttgga ctcgcttttc tgagaaagcg aggcttcaag ccaatgcaaa    660 

ctcctttttt catgagaaag gaaaccatgg gaaaatgtgc ccagttggcc caatttgacg    720 

aagagcttta caagctaaca ggcgatggag aggaaaagta tctcatcgct acatccgagc    780 

aaccgctgtg tgcgtatcat ctaggtgatc gaatttatcc tgctgaattg ccaattagat    840 

atgctggata ttccacctgc ttccggaaag aagctggttc acatggaagg gacacggctg    900 

gtatcttcag agtccaccaa ttcgaaaaga ttgagcaatt ctgtgttaca agtccaaatg    960 

acaatgaatc ctgggagatg catgaagaaa tgataaaaaa ttcagaagat ttctacaagg   1020 

agattggcct accgtaccaa cttgtctcca ttgtgtctgg tgctcttaat gatgctgcag   1080 

ctaagaagta tgatttagaa gcatggttcc ctgcatcaaa aacctatagg gaattagtgt   1140 

cctgctcaaa ttgcacagat tttcaagcaa ggagacttgg tataggttat ggccagaaaa   1200 

agaatgatag caatccaagc aattcgttca tatgttgaac tcaacattga ctgccactga   1260 

gaggaccctt tgctgtattt tggagaactt ccagaaggag aatggtgtcg aagttccaaa   1320 

agcattgcag ccttacatgg gtggaatcga tttccttcct ttcaagctgg atagcaaaca   1380 

agttgctcga ctccaaatca aataatccaa attcaaaggg agatgctatc tgagctagat   1440 

gaggaatcaa caaagatttt cttgctttca gacactactg gatgttattc atacttctaa   1500 

aaaatgcgtt tgttcagaac ttgtatcaat gatcatgatg ttacagtttt ggctctcatt   1560 

tgagtgtatt gattagcaca atgtctgacc atgtacttgc acagtgatat tccgtagaat   1620 

gtctggctat cttggacatg tgcgcttaat ttgccgtaaa agatgtattc attttcatgg   1680 

cctttagtgc ctatactaat ttgttgcata caaaaaaaaa aaaaaaaaaa aaaaaaaaaa   1740 

aaaaaaaaaa aaa                                                      1753 

 
           
             16  
             453  
             PRT  
             Oryza sativa  
             
               UNSURE  
               (390)  
               Xaa = any amino acid  
             
           
            16 

Met Leu Asp Ile Asn Leu Phe Arg Thr Glu Lys Gly Gly Asp Pro Glu 
  1               5                  10                  15 

Leu Ile Arg Arg Ser Gln Arg Asn Arg Ser Ala Ser Val Glu Leu Val 
             20                  25                  30 

Asp Glu Val Ile Ala Leu Asp Asp Gln Trp Arg Gln Arg Gln Phe Glu 
         35                  40                  45 

Leu Asp Lys Ile Arg Gln Glu Leu Asn Lys Thr Ser Lys Glu Ile Gly 
     50                  55                  60 

Lys Leu Lys Ala Lys Lys Glu Asp Ala Ser Ala Leu Ile Gln Ser Thr 
 65                  70                  75                  80 

Glu Glu Ile Lys Lys Arg Leu Ala Ala Lys Glu Thr Glu Val Gln Glu 
                 85                  90                  95 

Ala Lys Gly Thr Leu Asp Ala Lys Leu Val Thr Ile Gly Asn Ile Val 
            100                 105                 110 

His Glu Ser Val Pro Val Ser Asp Asp Glu Ala Asn Asn Leu Ile Val 
        115                 120                 125 

Arg Thr Trp Gly Glu Arg Arg Leu Glu Gly Asn Leu Lys Asn His Val 
    130                 135                 140 

Asp Leu Cys Lys Met Leu Asp Ile Val Ala Leu Glu Lys Gly Ala Asp 
145                 150                 155                 160 

Val Ala Gly Gly Arg Gly Tyr Tyr Leu Lys Asp Glu Gly Val Leu Leu 
                165                 170                 175 

Asn Leu Ala Leu Ile Asn Phe Gly Leu Ala Phe Leu Arg Lys Arg Gly 
            180                 185                 190 

Phe Lys Pro Met Gln Thr Pro Phe Phe Met Arg Lys Glu Thr Met Gly 
        195                 200                 205 

Lys Cys Ala Gln Leu Ala Gln Phe Asp Glu Glu Leu Tyr Lys Leu Thr 
    210                 215                 220 

Gly Asp Gly Glu Glu Lys Tyr Leu Ile Ala Thr Ser Glu Gln Pro Leu 
225                 230                 235                 240 

Cys Ala Tyr His Leu Gly Asp Arg Ile Tyr Pro Ala Glu Leu Pro Ile 
                245                 250                 255 

Arg Tyr Ala Gly Tyr Ser Thr Cys Phe Arg Lys Glu Ala Gly Ser His 
            260                 265                 270 

Gly Arg Asp Thr Ala Gly Ile Phe Arg Val His Gln Phe Glu Lys Ile 
        275                 280                 285 

Glu Gln Phe Cys Val Thr Ser Pro Asn Asp Asn Glu Ser Trp Glu Met 
    290                 295                 300 

His Glu Glu Met Ile Lys Asn Ser Glu Asp Phe Tyr Lys Glu Ile Gly 
305                 310                 315                 320 

Leu Pro Tyr Gln Leu Val Ser Ile Val Ser Gly Ala Leu Asn Asp Ala 
                325                 330                 335 

Ala Ala Lys Lys Tyr Asp Leu Glu Ala Trp Phe Pro Ala Ser Lys Thr 
            340                 345                 350 

Tyr Arg Glu Leu Val Ser Cys Ser Asn Cys Thr Asp Phe Gln Ala Arg 
        355                 360                 365 

Arg Leu Gly Ile Gly Tyr Gly Gln Lys Lys Asn Asp Ser Asn Pro Ser 
    370                 375                 380 

Asn Ser Phe Ile Cys Xaa Thr Gln His Xaa Leu Pro Leu Arg Gly Pro 
385                 390                 395                 400 

Phe Ala Val Phe Trp Arg Thr Ser Arg Arg Arg Met Val Ser Lys Phe 
                405                 410                 415 

Gln Lys His Cys Ser Leu Thr Trp Val Glu Ser Ile Ser Phe Leu Ser 
            420                 425                 430 

Ser Trp Ile Ala Asn Lys Leu Leu Asp Ser Lys Ser Asn Asn Pro Asn 
        435                 440                 445 

Ser Lys Gly Asp Ala 
    450 

 
           
             17  
             554  
             DNA  
             Glycine max  
           
            17 

cgcacgagat gatgctgcag caaagaagta tgatctagaa gcatggtttc cagcctctca     60 

agcttacaga gagctagtgt cctgttcaaa ctgtacagac tatcaggcca gaagattaga    120 

aattcgatat ggtcagaaaa agagcaatga gcaaatgaag caatatgttc acttgttgaa    180 

ctctactcta acggctactg agaggaccat ttgctgcata ctagagaaca accagaagga    240 

agatggggta gagataccag aagccctcag gccattcatg ggtggaaaga ctttcctacc    300 

tttcaagaac caaccatcta atgaagccaa agggaagaaa tcgaaggcct aattgcattt    360 

taagtcagtg atatttatga agagtttgtt cagagttgta ggttatgatg accccgatat    420 

tatccaccac acactgtttc cctttctcac taaatagtaa aatgattttt aggagcacgc    480 

accatttttg gtcaaagtac acagcatgac attttctgta attcattact ctaaaaaatt    540 

tgtgcttttt taac                                                      554 

 
           
             18  
             116  
             PRT  
             Glycine max  
           
            18 

Ala Arg Asp Asp Ala Ala Ala Lys Lys Tyr Asp Leu Glu Ala Trp Phe 
  1               5                  10                  15 

Pro Ala Ser Gln Ala Tyr Arg Glu Leu Val Ser Cys Ser Asn Cys Thr 
             20                  25                  30 

Asp Tyr Gln Ala Arg Arg Leu Glu Ile Arg Tyr Gly Gln Lys Lys Ser 
         35                  40                  45 

Asn Glu Gln Met Lys Gln Tyr Val His Leu Leu Asn Ser Thr Leu Thr 
     50                  55                  60 

Ala Thr Glu Arg Thr Ile Cys Cys Ile Leu Glu Asn Asn Gln Lys Glu 
 65                  70                  75                  80 

Asp Gly Val Glu Ile Pro Glu Ala Leu Arg Pro Phe Met Gly Gly Lys 
                 85                  90                  95 

Thr Phe Leu Pro Phe Lys Asn Gln Pro Ser Asn Glu Ala Lys Gly Lys 
            100                 105                 110 

Lys Ser Lys Ala 
        115 

 
           
             19  
             1719  
             DNA  
             Triticum aestivum  
           
            19 

ccgtcatcca cctctcgtca gaagaaccct ccctctccgc cctccgccgc cgccgccgcc     60 

gccgccaatc gcagcgcagc cgcctcgccc ccgtcgagct cgtccgcgga gaagttggcc    120 

tagccgcggt cgcacgagaa tatgctcgac atcaacctct tccgcaagga gaagggcggc    180 

gaccctgagc tcgtccgcca gtcgcagcgc agccgcttcg cccccgtcga gctcgtcgac    240 

gaggtcatcg tcctcgacga ggcgtggcgc cagaggcagt tcgagctcga caagatccgg    300 

caggagctca acaaaaccag caaggagatc ggcaagctca aggccaaaaa gcaggatgcg    360 

acggagctga tacagagcac ggaggagatt aagaagaggc tggccgccaa ggagacggac    420 

gtgcaggagg ccaagaccac cctcgatgcc aagctagtta ccatcggcaa cctcgtgcat    480 

gaatctgtgc ccatcagcaa cgacgaggca aacaatgcta ttgtgcggac atggggcgag    540 

aagagactgg aggagaaatt gaagaatcat gtggatcttt gcataatgct tgacatcgta    600 

tctttggata agggtgctga tgtagctggt ggaagaggtt tctttttgaa gggtgacggt    660 

gttctcctga accaggcgtt gataaatttt gggctatcat tcctgggaaa acgagaattt    720 

acaccaatgc aaactccttt tttcatgaga aaggagatca tggcaaaatg tgcccagttg    780 

gcccaatttg atgaggagct ctacaaagta acaggtgacg gagaggataa gtatctcata    840 

gcaacatcgg agcaaccgct atgtgcttat catctaggtg atcgaattta tcctgcagat    900 

ttgcctatca gatatgctgg gttctccacg tgcttccgga aagaagctgg ttcacacggg    960 

agggacacag ctggcatctt cagagtccac cagtttgaaa agatcgagca gttctgcgcc   1020 

acaggtccaa atgacaatgt atcctgggaa atgcatgagg agatgattaa aaatgcagaa   1080 

gatttttatc aggcgattgg gctaccatat caactagttt caattgtctc tggtgctctt   1140 

aatgatgctg cagctaagaa gtatgatttg gaagcatggt tccctgcatc aaaaaccttc   1200 

cgagaattag tgtcctgttc aaattgcaca gattatcagg caaggagact tggaataggc   1260 

tatggccaga aaaagaatga tgagcaatcg aagcagttcg ttcatatgtt gaattccacg   1320 

ctgactgcaa ctgagaggac actttgctgt attctggaga actaccagcg ggaaggtggt   1380 

gttgaagtgc cagaggtgtt gcggccattc atgcttggaa tagatttcct tcctttcaag   1440 

cggcctcttg ttgatagcaa acaagctgct gctgactcca aacccaataa gtctaaacca   1500 

aagggaaatg cagcttgaac tgaaaattgt ttccaggcag ataatgatgc acccttcctt   1560 

ttattaattt caagaatggt ctgtagcatg atgatgattt ggtctcccat tttggatggt   1620 

tttggttacg aagtattgca atccaggaca cataatttac cgcaaagtat attaatgttt   1680 

ttcatgacta aaaaaaaaaa aaaaaaaact cgagactag                          1719 

 
           
             20  
             458  
             PRT  
             Triticum aestivum  
           
            20 

Met Leu Asp Ile Asn Leu Phe Arg Lys Glu Lys Gly Gly Asp Pro Glu 
  1               5                  10                  15 

Leu Val Arg Gln Ser Gln Arg Ser Arg Phe Ala Pro Val Glu Leu Val 
             20                  25                  30 

Asp Glu Val Ile Val Leu Asp Glu Ala Trp Arg Gln Arg Gln Phe Glu 
         35                  40                  45 

Leu Asp Lys Ile Arg Gln Glu Leu Asn Lys Thr Ser Lys Glu Ile Gly 
     50                  55                  60 

Lys Leu Lys Ala Lys Lys Gln Asp Ala Thr Glu Leu Ile Gln Ser Thr 
 65                  70                  75                  80 

Glu Glu Ile Lys Lys Arg Leu Ala Ala Lys Glu Thr Asp Val Gln Glu 
                 85                  90                  95 

Ala Lys Thr Thr Leu Asp Ala Lys Leu Val Thr Ile Gly Asn Leu Val 
            100                 105                 110 

His Glu Ser Val Pro Ile Ser Asn Asp Glu Ala Asn Asn Ala Ile Val 
        115                 120                 125 

Arg Thr Trp Gly Glu Lys Arg Leu Glu Glu Lys Leu Lys Asn His Val 
    130                 135                 140 

Asp Leu Cys Ile Met Leu Asp Ile Val Ser Leu Asp Lys Gly Ala Asp 
145                 150                 155                 160 

Val Ala Gly Gly Arg Gly Phe Phe Leu Lys Gly Asp Gly Val Leu Leu 
                165                 170                 175 

Asn Gln Ala Leu Ile Asn Phe Gly Leu Ser Phe Leu Gly Lys Arg Glu 
            180                 185                 190 

Phe Thr Pro Met Gln Thr Pro Phe Phe Met Arg Lys Glu Ile Met Ala 
        195                 200                 205 

Lys Cys Ala Gln Leu Ala Gln Phe Asp Glu Glu Leu Tyr Lys Val Thr 
    210                 215                 220 

Gly Asp Gly Glu Asp Lys Tyr Leu Ile Ala Thr Ser Glu Gln Pro Leu 
225                 230                 235                 240 

Cys Ala Tyr His Leu Gly Asp Arg Ile Tyr Pro Ala Asp Leu Pro Ile 
                245                 250                 255 

Arg Tyr Ala Gly Phe Ser Thr Cys Phe Arg Lys Glu Ala Gly Ser His 
            260                 265                 270 

Gly Arg Asp Thr Ala Gly Ile Phe Arg Val His Gln Phe Glu Lys Ile 
        275                 280                 285 

Glu Gln Phe Cys Ala Thr Gly Pro Asn Asp Asn Val Ser Trp Glu Met 
    290                 295                 300 

His Glu Glu Met Ile Lys Asn Ala Glu Asp Phe Tyr Gln Ala Ile Gly 
305                 310                 315                 320 

Leu Pro Tyr Gln Leu Val Ser Ile Val Ser Gly Ala Leu Asn Asp Ala 
                325                 330                 335 

Ala Ala Lys Lys Tyr Asp Leu Glu Ala Trp Phe Pro Ala Ser Lys Thr 
            340                 345                 350 

Phe Arg Glu Leu Val Ser Cys Ser Asn Cys Thr Asp Tyr Gln Ala Arg 
        355                 360                 365 

Arg Leu Gly Ile Gly Tyr Gly Gln Lys Lys Asn Asp Glu Gln Ser Lys 
    370                 375                 380 

Gln Phe Val His Met Leu Asn Ser Thr Leu Thr Ala Thr Glu Arg Thr 
385                 390                 395                 400 

Leu Cys Cys Ile Leu Glu Asn Tyr Gln Arg Glu Gly Gly Val Glu Val 
                405                 410                 415 

Pro Glu Val Leu Arg Pro Phe Met Leu Gly Ile Asp Phe Leu Pro Phe 
            420                 425                 430 

Lys Arg Pro Leu Val Asp Ser Lys Gln Ala Ala Ala Asp Ser Lys Pro 
        435                 440                 445 

Asn Lys Ser Lys Pro Lys Gly Asn Ala Ala 
    450                 455 

 
           
             21  
             2445  
             DNA  
             Zea mays  
           
            21 

ggcgaccggc cagcgccata tcccgcgccg ccgccgccgc cgccgccgcc gccaacgctt     60 

aatgctagtg ttcctccggc gaggcctctt gctcgtccgg cagcccacca cccgcgtcct    120 

tgccaaaccg tctctccgcc ctgcttgtct cttcgtccac cacttcgccg tcgacacgat    180 

gggtgagggt tctgctgctg gtaaggacgc gaaggggaag gggaagggga aggggaagac    240 

caaggccgcc gccgcggatt cggccctggt cgttcgcgac gactcctacc tagaggcggt    300 

cactcagaag aggattcgct tcttcgagga gatccaggca aggcaagccg tcgagcggct    360 

gaatatcggc ggcgaagtta tcaaggtaac tttgcctgat ggcgctatca aggagggtaa    420 

gaaatggata acaaccccaa tggatattgc taaggagata tcaagtggat ttgcagctag    480 

ttgtttgata gctcaagtgg acgaaacact ctgggacatg gggaggccac tcgaaggtga    540 

ttgtaaattg caaatgttca agtttgatac caatgaaggt cgtgacacct tctggcactc    600 

aagtgctcat attcttggag aatctattga gagagcatat ggatgcaagc tgtgtattgg    660 

gccttgcacc acaagagggg agggtttcta ctatgatgct tactacaatg atcagacatt    720 

gaatgaggag cactttggta tcattgaaaa ccaagctaaa aaggctgttg cggaaaagca    780 

accgtttgaa cgcattgagg tcagcagggc agaagctctt gaaatgtttg ctgagaatga    840 

attcaaggtt gaaatcatta atgagttgcc tgaggacaag accattactg tatacaggtg    900 

tggtccttta gttgacctat gccgtgggcc acacatccca aatacttcct ttgtcaaagc    960 

tttcgcttgt ctgaaggctt catcatcata ttggagagga aaagttgacc gcgaaagcct   1020 

gcagagagta tatggaattt ctttccctga ttctcgacgt ctcacggaat ataaacattt   1080 

tctagaggaa gctaagaaac gtgatcatag gatattaggg aaagcacagg aactcttctt   1140 

tttccatgaa cttagccctg gaagctgctt cttccttcca catggtgcca ggatatataa   1200 

caaactgatg gacttcatgc gacaacagta cagagataga ggataccaag aggtgttgag   1260 

cccaaatatt tacaatatgc aactatggga aacttctgga cacgccgcaa actataagga   1320 

gaacatgttt gtttttgaga tcgagaaaca ggaatttgga cttaagccaa tgaattgtcc   1380 

aggacactgt ctaatgtttg ctaatagggt tcggtcgtac agagagttgc ctcttcgcat   1440 

ggctgatttt ggagtgcttc atagaaatga gcttagtggt gctcttacag gtttgacacg   1500 

tgttagaaga ttccagcagg atgatgctca tatcttctgc agagaagacc aaatcaagga   1560 

tgaagttaag gctgttttgg aattcatcaa ttatgtttat gagatatttg gcttcaaata   1620 

tgagttggag ttgtctacga gaccagagaa gtatctaggt gaagttgaaa cttggaacaa   1680 

agctgaacaa caattgacag aagctttaaa tgagtttggg aagccatgga agattaatga   1740 

aggggatggt gctttctacg gcccaaaaat tgatattggt gtgtttgatg cccttaagag   1800 

gaaatttcag tgtgcaaccc tacagctgga ttttcagctg cccattcggt tcaagctggc   1860 

ttattctgct gaggatgaag ccaaaattga aaggcctgtg atgatacaca gggcaatcct   1920 

aggttcggtt gaaaggatgc ttgccattct tttggagcat tacaatggta aatggccctt   1980 

atggctaagc cctcgccagg ccattgtttg ctcggtatct tctggttcag tggaatatgc   2040 

gaaacaggtt cttgccactc tacatgaagc tggttttcat gttgatattg acgcgagtga   2100 

caggacaata caaaagaagg tacgggaagc tcaactggcc caattcaact acattcttgt   2160 

cgtaggtgca caagaggccg agactggaaa tatatgcgtt agggtaagag acaatgctga   2220 

cctggtcaca acgagtgtag atggcttcat cacacgtttg agggacgaaa tcgcagcctt   2280 

caaatgattt tgatgctgca taatttccta ctactcgttt gtgattttga cgagttttta   2340 

gtgaccagca tcgagttcct cgtgttactg ttcttgtttg tatgaagcta aaaggttgtc   2400 

tttttgttat taattacaga tgcgaagtta aatactgccg ctagt                   2445 

 
           
             22  
             741  
             PRT  
             Zea mays  
           
            22 

Met Leu Val Phe Leu Arg Arg Gly Leu Leu Leu Val Arg Gln Pro Thr 
  1               5                  10                  15 

Thr Arg Val Leu Ala Lys Pro Ser Leu Arg Pro Ala Cys Leu Phe Val 
             20                  25                  30 

His His Phe Ala Val Asp Thr Met Gly Glu Gly Ser Ala Ala Gly Lys 
         35                  40                  45 

Asp Ala Lys Gly Lys Gly Lys Gly Lys Gly Lys Thr Lys Ala Ala Ala 
     50                  55                  60 

Ala Asp Ser Ala Leu Val Val Arg Asp Asp Ser Tyr Leu Glu Ala Val 
 65                  70                  75                  80 

Thr Gln Lys Arg Ile Arg Phe Phe Glu Glu Ile Gln Ala Arg Gln Ala 
                 85                  90                  95 

Val Glu Arg Leu Asn Ile Gly Gly Glu Val Ile Lys Val Thr Leu Pro 
            100                 105                 110 

Asp Gly Ala Ile Lys Glu Gly Lys Lys Trp Ile Thr Thr Pro Met Asp 
        115                 120                 125 

Ile Ala Lys Glu Ile Ser Ser Gly Phe Ala Ala Ser Cys Leu Ile Ala 
    130                 135                 140 

Gln Val Asp Glu Thr Leu Trp Asp Met Gly Arg Pro Leu Glu Gly Asp 
145                 150                 155                 160 

Cys Lys Leu Gln Met Phe Lys Phe Asp Thr Asn Glu Gly Arg Asp Thr 
                165                 170                 175 

Phe Trp His Ser Ser Ala His Ile Leu Gly Glu Ser Ile Glu Arg Ala 
            180                 185                 190 

Tyr Gly Cys Lys Leu Cys Ile Gly Pro Cys Thr Thr Arg Gly Glu Gly 
        195                 200                 205 

Phe Tyr Tyr Asp Ala Tyr Tyr Asn Asp Gln Thr Leu Asn Glu Glu His 
    210                 215                 220 

Phe Gly Ile Ile Glu Asn Gln Ala Lys Lys Ala Val Ala Glu Lys Gln 
225                 230                 235                 240 

Pro Phe Glu Arg Ile Glu Val Ser Arg Ala Glu Ala Leu Glu Met Phe 
                245                 250                 255 

Ala Glu Asn Glu Phe Lys Val Glu Ile Ile Asn Glu Leu Pro Glu Asp 
            260                 265                 270 

Lys Thr Ile Thr Val Tyr Arg Cys Gly Pro Leu Val Asp Leu Cys Arg 
        275                 280                 285 

Gly Pro His Ile Pro Asn Thr Ser Phe Val Lys Ala Phe Ala Cys Leu 
    290                 295                 300 

Lys Ala Ser Ser Ser Tyr Trp Arg Gly Lys Val Asp Arg Glu Ser Leu 
305                 310                 315                 320 

Gln Arg Val Tyr Gly Ile Ser Phe Pro Asp Ser Arg Arg Leu Thr Glu 
                325                 330                 335 

Tyr Lys His Phe Leu Glu Glu Ala Lys Lys Arg Asp His Arg Ile Leu 
            340                 345                 350 

Gly Lys Ala Gln Glu Leu Phe Phe Phe His Glu Leu Ser Pro Gly Ser 
        355                 360                 365 

Cys Phe Phe Leu Pro His Gly Ala Arg Ile Tyr Asn Lys Leu Met Asp 
    370                 375                 380 

Phe Met Arg Gln Gln Tyr Arg Asp Arg Gly Tyr Gln Glu Val Leu Ser 
385                 390                 395                 400 

Pro Asn Ile Tyr Asn Met Gln Leu Trp Glu Thr Ser Gly His Ala Ala 
                405                 410                 415 

Asn Tyr Lys Glu Asn Met Phe Val Phe Glu Ile Glu Lys Gln Glu Phe 
            420                 425                 430 

Gly Leu Lys Pro Met Asn Cys Pro Gly His Cys Leu Met Phe Ala Asn 
        435                 440                 445 

Arg Val Arg Ser Tyr Arg Glu Leu Pro Leu Arg Met Ala Asp Phe Gly 
    450                 455                 460 

Val Leu His Arg Asn Glu Leu Ser Gly Ala Leu Thr Gly Leu Thr Arg 
465                 470                 475                 480 

Val Arg Arg Phe Gln Gln Asp Asp Ala His Ile Phe Cys Arg Glu Asp 
                485                 490                 495 

Gln Ile Lys Asp Glu Val Lys Ala Val Leu Glu Phe Ile Asn Tyr Val 
            500                 505                 510 

Tyr Glu Ile Phe Gly Phe Lys Tyr Glu Leu Glu Leu Ser Thr Arg Pro 
        515                 520                 525 

Glu Lys Tyr Leu Gly Glu Val Glu Thr Trp Asn Lys Ala Glu Gln Gln 
    530                 535                 540 

Leu Thr Glu Ala Leu Asn Glu Phe Gly Lys Pro Trp Lys Ile Asn Glu 
545                 550                 555                 560 

Gly Asp Gly Ala Phe Tyr Gly Pro Lys Ile Asp Ile Gly Val Phe Asp 
                565                 570                 575 

Ala Leu Lys Arg Lys Phe Gln Cys Ala Thr Leu Gln Leu Asp Phe Gln 
            580                 585                 590 

Leu Pro Ile Arg Phe Lys Leu Ala Tyr Ser Ala Glu Asp Glu Ala Lys 
        595                 600                 605 

Ile Glu Arg Pro Val Met Ile His Arg Ala Ile Leu Gly Ser Val Glu 
    610                 615                 620 

Arg Met Leu Ala Ile Leu Leu Glu His Tyr Asn Gly Lys Trp Pro Leu 
625                 630                 635                 640 

Trp Leu Ser Pro Arg Gln Ala Ile Val Cys Ser Val Ser Ser Gly Ser 
                645                 650                 655 

Val Glu Tyr Ala Lys Gln Val Leu Ala Thr Leu His Glu Ala Gly Phe 
            660                 665                 670 

His Val Asp Ile Asp Ala Ser Asp Arg Thr Ile Gln Lys Lys Val Arg 
        675                 680                 685 

Glu Ala Gln Leu Ala Gln Phe Asn Tyr Ile Leu Val Val Gly Ala Gln 
    690                 695                 700 

Glu Ala Glu Thr Gly Asn Ile Cys Val Arg Val Arg Asp Asn Ala Asp 
705                 710                 715                 720 

Leu Val Thr Thr Ser Val Asp Gly Phe Ile Thr Arg Leu Arg Asp Glu 
                725                 730                 735 

Ile Ala Ala Phe Lys 
            740 

 
           
             23  
             1758  
             DNA  
             Oryza sativa  
           
            23 

gcacgagatt atgatgccta ctacaatgat ctgacattga atgagacaca ttttggtatc     60 

attgatgccc aagcacagaa agctgttgcg gaaaaacaac catttgaacg aattgaggtc    120 

tccagggcag aggcccttga aatgttcgca gaaaataaat ttaaggttga aatcattaat    180 

gagttgcctg aagacaagac cattacagta tacagatgtg gtcctctagt tgacctttgc    240 

cgtgggccac acatccccaa tacttccttt gttaaagctt ttgcttgtct taaggcatca    300 

tcgtcgtatt ggagagggaa agcagatcga gagagcctgc agagagtata tggaatttct    360 

tttcctgatt ctaaacgtct caaggaatat aaacatctgc tagaagaggc taagaagcgt    420 

gatcataggc tattaggaca gacccaggat ctcttctttt tccatcaact tagtccagga    480 

agctgcttct tccttccaca tggtgctata atatacaaca aattgatgga ttttttgcga    540 

cagcaataca gagatagagg atatcaagag gttttgagcc caaatattta caatatgcaa    600 

ctctgggaaa cctctggaca tgctgcaaac tacaaggaga atatgtttgt ttttgagatt    660 

gagaaacagg aatttggtct caagccaatg aattgtcctg gccattgcct aatgtttgag    720 

cacagggttc gttcatacag agaattgcct ctccggatgg ctgattttgg agtccttcac    780 

aggaatgagc ttagtggtgc acttacaggt ttgacacgtg ttagaagatt ccagcaggat    840 

gatgcccata ttttttgcag agaaagccaa atcaaggacg aagttaaggc tgttttggac    900 

ttcatcaatt atgtttacga gatatttgga tttaaatatg aattggagct atcaacgaga    960 

ccagaaaagt acttaggtga tattgaaacc tggaacaaag cagagcaaca gctgacagaa   1020 

gccttaaatg agtttggaaa gccatggcag atcaatgaag gtgatggtgc cttctatggt   1080 

ccaaaaattg atattggtgt gtttgatgcc ctcaagagga aatttcagtg tgcaactcta   1140 

cagctcgatt ttcagctgcc cctacgcttc aagctgactt actctgcaga ggatgaagcc   1200 

aaacttgaga ggcctgtgat gattcacagg gcaatcctag gttctgttga aaggatgttt   1260 

gctattcttt tggagcatta caatggtaaa tggcccttgt ggttgagtcc tcgccaagcc   1320 

attgtttgct ccatatcttc caattcagtg gaatacgcta aacaggtccg tgctaggata   1380 

catgaagctg gttttcatgt agccatcgat gagacagaca ggacaataca gaagaaggta   1440 

cgggaggctc aattagccca attcaactac attcttgtcg ttggtgcaca agaagcagag   1500 

actggacagg tcagcgtcag ggtaagggac aaagctgaac tagccacagt gagcattgat   1560 

gacatcatca cacgttttaa ggaggaagta gcagcttaca aatgattttg atttcacacc   1620 

cttttgctaa gaatttactc caaatttgtg attttgatgg tgtagcgggc agtgtaatct   1680 

tgctatttta tttcttgaca aaagtacatc tgattgtctt ttcttaataa cgaaagtgtg   1740 

ctattcttca tcagcgac                                                 1758 

 
           
             24  
             530  
             PRT  
             Oryza sativa  
           
            24 

Tyr Asp Ala Tyr Tyr Asn Asp Leu Thr Leu Asn Glu Thr His Phe Gly 
  1               5                  10                  15 

Ile Ile Asp Ala Gln Ala Gln Lys Ala Val Ala Glu Lys Gln Pro Phe 
             20                  25                  30 

Glu Arg Ile Glu Val Ser Arg Ala Glu Ala Leu Glu Met Phe Ala Glu 
         35                  40                  45 

Asn Lys Phe Lys Val Glu Ile Ile Asn Glu Leu Pro Glu Asp Lys Thr 
     50                  55                  60 

Ile Thr Val Tyr Arg Cys Gly Pro Leu Val Asp Leu Cys Arg Gly Pro 
 65                  70                  75                  80 

His Ile Pro Asn Thr Ser Phe Val Lys Ala Phe Ala Cys Leu Lys Ala 
                 85                  90                  95 

Ser Ser Ser Tyr Trp Arg Gly Lys Ala Asp Arg Glu Ser Leu Gln Arg 
            100                 105                 110 

Val Tyr Gly Ile Ser Phe Pro Asp Ser Lys Arg Leu Lys Glu Tyr Lys 
        115                 120                 125 

His Leu Leu Glu Glu Ala Lys Lys Arg Asp His Arg Leu Leu Gly Gln 
    130                 135                 140 

Thr Gln Asp Leu Phe Phe Phe His Gln Leu Ser Pro Gly Ser Cys Phe 
145                 150                 155                 160 

Phe Leu Pro His Gly Ala Ile Ile Tyr Asn Lys Leu Met Asp Phe Leu 
                165                 170                 175 

Arg Gln Gln Tyr Arg Asp Arg Gly Tyr Gln Glu Val Leu Ser Pro Asn 
            180                 185                 190 

Ile Tyr Asn Met Gln Leu Trp Glu Thr Ser Gly His Ala Ala Asn Tyr 
        195                 200                 205 

Lys Glu Asn Met Phe Val Phe Glu Ile Glu Lys Gln Glu Phe Gly Leu 
    210                 215                 220 

Lys Pro Met Asn Cys Pro Gly His Cys Leu Met Phe Glu His Arg Val 
225                 230                 235                 240 

Arg Ser Tyr Arg Glu Leu Pro Leu Arg Met Ala Asp Phe Gly Val Leu 
                245                 250                 255 

His Arg Asn Glu Leu Ser Gly Ala Leu Thr Gly Leu Thr Arg Val Arg 
            260                 265                 270 

Arg Phe Gln Gln Asp Asp Ala His Ile Phe Cys Arg Glu Ser Gln Ile 
        275                 280                 285 

Lys Asp Glu Val Lys Ala Val Leu Asp Phe Ile Asn Tyr Val Tyr Glu 
    290                 295                 300 

Ile Phe Gly Phe Lys Tyr Glu Leu Glu Leu Ser Thr Arg Pro Glu Lys 
305                 310                 315                 320 

Tyr Leu Gly Asp Ile Glu Thr Trp Asn Lys Ala Glu Gln Gln Leu Thr 
                325                 330                 335 

Glu Ala Leu Asn Glu Phe Gly Lys Pro Trp Gln Ile Asn Glu Gly Asp 
            340                 345                 350 

Gly Ala Phe Tyr Gly Pro Lys Ile Asp Ile Gly Val Phe Asp Ala Leu 
        355                 360                 365 

Lys Arg Lys Phe Gln Cys Ala Thr Leu Gln Leu Asp Phe Gln Leu Pro 
    370                 375                 380 

Leu Arg Phe Lys Leu Thr Tyr Ser Ala Glu Asp Glu Ala Lys Leu Glu 
385                 390                 395                 400 

Arg Pro Val Met Ile His Arg Ala Ile Leu Gly Ser Val Glu Arg Met 
                405                 410                 415 

Phe Ala Ile Leu Leu Glu His Tyr Asn Gly Lys Trp Pro Leu Trp Leu 
            420                 425                 430 

Ser Pro Arg Gln Ala Ile Val Cys Ser Ile Ser Ser Asn Ser Val Glu 
        435                 440                 445 

Tyr Ala Lys Gln Val Arg Ala Arg Ile His Glu Ala Gly Phe His Val 
    450                 455                 460 

Ala Ile Asp Glu Thr Asp Arg Thr Ile Gln Lys Lys Val Arg Glu Ala 
465                 470                 475                 480 

Gln Leu Ala Gln Phe Asn Tyr Ile Leu Val Val Gly Ala Gln Glu Ala 
                485                 490                 495 

Glu Thr Gly Gln Val Ser Val Arg Val Arg Asp Lys Ala Glu Leu Ala 
            500                 505                 510 

Thr Val Ser Ile Asp Asp Ile Ile Thr Arg Phe Lys Glu Glu Val Ala 
        515                 520                 525 

Ala Tyr 
    530 

 
           
             25  
             2380  
             DNA  
             Glycine max  
           
            25 

gcacgagaca ctatgctatg ctatgctctc taatccgttt ccgccgttac gctccttcct     60 

accgcactct ccactctctc tttccgacga ttaaccgttt ctcctcctcc gtctcctccg    120 

cctccgccgc catggttgct cacgcgaagg acgaggcgta cctcagcgcg acgattccga    180 

aacgcatccg tctcttcgag accatcctgg cggagcagca cactcagcgc ctctcgctct    240 

ccccggatcc tatcaaggtt actctccccg acggcagcgt caaggaggcg aagaagtggc    300 

atacgacgcc gcttgatgtt gcgcgtgaaa tctcgaagaa tttggccaac agcgcgctca    360 

tcgcgaaggt caatggcgtg ctctgggaca tgactcgccc tctcgaggac gattgccagc    420 

tccagatctt caagttcgac gacgacgaag gccgcgacac cttctggcac tccagcgccc    480 

acattctcgg ccagtcactt gagacggagt atggatgcaa gctctgcatt gggccttgca    540 

ctacaagagg agagggattc tattatgatg cattttacgg ggagttgggt ctcaatgacg    600 

atcactttaa gcagattgag gctggagcat tgaaggctgt tgcggaaaag caaccctttg    660 

agcgtattga agttacacgt gatcaggcac ttgagatgtt ttcagataat aagtttaagg    720 

ttgagattat caatgatttg cctgccgaca aaactatcac agtatacaga tgtggcccct    780 

tggttgattt gtgtcgtgga ccccatatac ctaatacatc ctttgtcaaa gcaattgcgt    840 

gcttaaaggc ttcatcagca tattggaggg gggacaaaga tcgggaaagt ttacaaagag    900 

tttatggcat atcttatcct gatcagaaaa gtctaaagga atacttgcat cggctggagg    960 

aggctaaaaa gtatgatcac aggattttgg gtgtgaaaca ggagcttatt cttcatcatg   1020 

aatggagccc gggaagctgg ttttttcttc cgcaaggcac tcggatctac aacaaactca   1080 

tggacttcat tcggaatcag tacagagaca ggggctatca agaggtcata tctcccaatg   1140 

tatttaacat ggaactgtgg gtgcaatctg gtcatgctgc aaattatagg gaggatatgt   1200 

ttatcttaga ggttgacaaa caagagtttg ggttgaaacc aatgaattgc ccagggcact   1260 

gcctgatgtt taaacacagg gttcgatcat atagagaact tcctcttcgt ttcgctgatt   1320 

ttggggtttt gcatcggaat gaggctagtg gcgccctgag tggattaaca cgtgttagga   1380 

gattccagca ggacgatgca catattttct gcagggagtc ccagataaag gatgaagtga   1440 

ggaacagctt gaatttcatc aattatgtct ataagatatt tggtttcaca tatgagctga   1500 

agctttcaac gaggccagaa aaatacctag gagatattgc aacttgggac aaagctgaaa   1560 

gtgctcttaa agaagcttta gatgattttg gcaagccttg gcagttgaat gaaggggatg   1620 

gtgcattcta tggaccaaag atagacatca gtgtatctga tgcattgggt aggaaattcc   1680 

agtgtgcaac tttgcagctt gacttccagc ttcctgatcg ttttaagttg gaattctcag   1740 

ctgaggatga agccaaaatt gagagacctg taatgataca cagagccatt ctaggatctg   1800 

ttgaacgcat gtttgccata cttttagagc actacaaggg taaatggcct ttctggctca   1860 

gtcctcgtca agcaattgta tgccctgtgt ctgaaaagtc acaagcttat gcattacagg   1920 

tgcgagatca gatccaccaa gcagggtatt acgttgatgc tgatacaact gataggaaga   1980 

ttcaaaagaa ggtgcgagaa gcacaattag cacaatacaa ctacatcttg gttgttggag   2040 

aggaggaagc taatacagga caggtgagtg tacgagttag agacttggca gaacataagg   2100 

ttatgagtat tgagaagcta cttgaacatt tcagagacaa agctgcagct ttcgaatgat   2160 

actttgcatg tgaaaactgt cgaagaaaat tttcagcccc aaatacctta gttttacaca   2220 

gttgtgtgcg cattttgatt ttcaacttaa gcaatttatc ctgattttat ttatgatttg   2280 

aatgatcact gtatttcgca ctagaaacat aatgtgaatc ttggtcatac ctggagcgca   2340 

ctctggttga tctttatatc aaaaaaaaaa aaaaaaaaaa                         2380 

 
           
             26  
             698  
             PRT  
             Glycine max  
           
            26 

Tyr Arg Thr Leu His Ser Leu Phe Pro Thr Ile Asn Arg Phe Ser Ser 
  1               5                  10                  15 

Ser Val Ser Ser Ala Ser Ala Ala Met Val Ala His Ala Lys Asp Glu 
             20                  25                  30 

Ala Tyr Leu Ser Ala Thr Ile Pro Lys Arg Ile Arg Leu Phe Glu Thr 
         35                  40                  45 

Ile Leu Ala Glu Gln His Thr Gln Arg Leu Ser Leu Ser Pro Asp Pro 
     50                  55                  60 

Ile Lys Val Thr Leu Pro Asp Gly Ser Val Lys Glu Ala Lys Lys Trp 
 65                  70                  75                  80 

His Thr Thr Pro Leu Asp Val Ala Arg Glu Ile Ser Lys Asn Leu Ala 
                 85                  90                  95 

Asn Ser Ala Leu Ile Ala Lys Val Asn Gly Val Leu Trp Asp Met Thr 
            100                 105                 110 

Arg Pro Leu Glu Asp Asp Cys Gln Leu Gln Ile Phe Lys Phe Asp Asp 
        115                 120                 125 

Asp Glu Gly Arg Asp Thr Phe Trp His Ser Ser Ala His Ile Leu Gly 
    130                 135                 140 

Gln Ser Leu Glu Thr Glu Tyr Gly Cys Lys Leu Cys Ile Gly Pro Cys 
145                 150                 155                 160 

Thr Thr Arg Gly Glu Gly Phe Tyr Tyr Asp Ala Phe Tyr Gly Glu Leu 
                165                 170                 175 

Gly Leu Asn Asp Asp His Phe Lys Gln Ile Glu Ala Gly Ala Leu Lys 
            180                 185                 190 

Ala Val Ala Glu Lys Gln Pro Phe Glu Arg Ile Glu Val Thr Arg Asp 
        195                 200                 205 

Gln Ala Leu Glu Met Phe Ser Asp Asn Lys Phe Lys Val Glu Ile Ile 
    210                 215                 220 

Asn Asp Leu Pro Ala Asp Lys Thr Ile Thr Val Tyr Arg Cys Gly Pro 
225                 230                 235                 240 

Leu Val Asp Leu Cys Arg Gly Pro His Ile Pro Asn Thr Ser Phe Val 
                245                 250                 255 

Lys Ala Ile Ala Cys Leu Lys Ala Ser Ser Ala Tyr Trp Arg Gly Asp 
            260                 265                 270 

Lys Asp Arg Glu Ser Leu Gln Arg Val Tyr Gly Ile Ser Tyr Pro Asp 
        275                 280                 285 

Gln Lys Ser Leu Lys Glu Tyr Leu His Arg Leu Glu Glu Ala Lys Lys 
    290                 295                 300 

Tyr Asp His Arg Ile Leu Gly Val Lys Gln Glu Leu Ile Leu His His 
305                 310                 315                 320 

Glu Trp Ser Pro Gly Ser Trp Phe Phe Leu Pro Gln Gly Thr Arg Ile 
                325                 330                 335 

Tyr Asn Lys Leu Met Asp Phe Ile Arg Asn Gln Tyr Arg Asp Arg Gly 
            340                 345                 350 

Tyr Gln Glu Val Ile Ser Pro Asn Val Phe Asn Met Glu Leu Trp Val 
        355                 360                 365 

Gln Ser Gly His Ala Ala Asn Tyr Arg Glu Asp Met Phe Ile Leu Glu 
    370                 375                 380 

Val Asp Lys Gln Glu Phe Gly Leu Lys Pro Met Asn Cys Pro Gly His 
385                 390                 395                 400 

Cys Leu Met Phe Lys His Arg Val Arg Ser Tyr Arg Glu Leu Pro Leu 
                405                 410                 415 

Arg Phe Ala Asp Phe Gly Val Leu His Arg Asn Glu Ala Ser Gly Ala 
            420                 425                 430 

Leu Ser Gly Leu Thr Arg Val Arg Arg Phe Gln Gln Asp Asp Ala His 
        435                 440                 445 

Ile Phe Cys Arg Glu Ser Gln Ile Lys Asp Glu Val Arg Asn Ser Leu 
    450                 455                 460 

Asn Phe Ile Asn Tyr Val Tyr Lys Ile Phe Gly Phe Thr Tyr Glu Leu 
465                 470                 475                 480 

Lys Leu Ser Thr Arg Pro Glu Lys Tyr Leu Gly Asp Ile Ala Thr Trp 
                485                 490                 495 

Asp Lys Ala Glu Ser Ala Leu Lys Glu Ala Leu Asp Asp Phe Gly Lys 
            500                 505                 510 

Pro Trp Gln Leu Asn Glu Gly Asp Gly Ala Phe Tyr Gly Pro Lys Ile 
        515                 520                 525 

Asp Ile Ser Val Ser Asp Ala Leu Gly Arg Lys Phe Gln Cys Ala Thr 
    530                 535                 540 

Leu Gln Leu Asp Phe Gln Leu Pro Asp Arg Phe Lys Leu Glu Phe Ser 
545                 550                 555                 560 

Ala Glu Asp Glu Ala Lys Ile Glu Arg Pro Val Met Ile His Arg Ala 
                565                 570                 575 

Ile Leu Gly Ser Val Glu Arg Met Phe Ala Ile Leu Leu Glu His Tyr 
            580                 585                 590 

Lys Gly Lys Trp Pro Phe Trp Leu Ser Pro Arg Gln Ala Ile Val Cys 
        595                 600                 605 

Pro Val Ser Glu Lys Ser Gln Ala Tyr Ala Leu Gln Val Arg Asp Gln 
    610                 615                 620 

Ile His Gln Ala Gly Tyr Tyr Val Asp Ala Asp Thr Thr Asp Arg Lys 
625                 630                 635                 640 

Ile Gln Lys Lys Val Arg Glu Ala Gln Leu Ala Gln Tyr Asn Tyr Ile 
                645                 650                 655 

Leu Val Val Gly Glu Glu Glu Ala Asn Thr Gly Gln Val Ser Val Arg 
            660                 665                 670 

Val Arg Asp Leu Ala Glu His Lys Val Met Ser Ile Glu Lys Leu Leu 
        675                 680                 685 

Glu His Phe Arg Asp Lys Ala Ala Ala Phe 
    690                 695 

 
           
             27  
             1677  
             DNA  
             Triticum aestivum  
           
            27 

cggaaaatga atttaaggtt gaaataatta acgaattgcc cgaggacaag accattacag     60 

tatacagatg tggtcctttg gtcgacctct gccgtggccc gcacatccca aatacttcct    120 

ttgttaaagc tttcgcttgc ctcaaggctt cagcatcata ctggagagga aaagcagacc    180 

gtgagagcct gcagagagta tatggaatct ccttccctga ttctaaacgt ctcaaggaat    240 

atcaacatat gatagaggaa gctaagaaac gcgatcatag gttactaggg cagtcccaga    300 

aactcttctt tttccatcca cttagcccag gtagctgctt cttccttcca aatggcgcta    360 

taatatataa caaattgatg gattttttgc gcaaggagta tagagagaga ggctaccaag    420 

aggttctgag tccaaatatt tacaacatgc aactttggga aacctctgga catgctgcaa    480 

actacaagga caacatgttt gtttttgaga tcgagaaaca agaatttggc cttaagccaa    540 

tgaattgtcc tggccattgc ctaatgtttg gacacgaggt tcgatcgtat agagagttgc    600 

ctctccgcat ggctgatttt ggagttctgc acagaaatga acttagtggt gcacttacag    660 

gtttgacacg tgtcagaaga ttccaacagg acgatgccca tattttttgc atggagagcc    720 

aaatcaagga tgaagttcgg gcttgcttgg agttcattga ttatgtttat aaaatatttg    780 

ggtttgaata tgagctggag ttatcaacga gaccagagaa gtatttaggt gacattgaga    840 

cctggaacaa agcagagcaa caactgacag aagcattgaa tgagtttggg aagccatgga    900 

agataaatga agcagatggt gctttctatg gcccgaaaat agatattggt gtgtttgatg    960 

ccctcaagag gaaatttcag tgtgcaactc tacagctcga ttttcagctg ccacttcgct   1020 

tcaagttgac ttattctgca gaggatgaag ccaagcttga gaggcctgta atgatacaca   1080 

gggcaatact aggatcagtt gaaaggatgt ttgccattct tttggagcac tataatggta   1140 

aatggccgtt gtggttaagc ccccgacaag ccattgtttg ctgtgtatct gccaattcac   1200 

taacatatgc aaaagaggtt catgctcaga tacgtgcagc tggttttcat gttgacattg   1260 

acatgactga tagaacaatt caaaagaagg tgcgggaggc tcagttagcc caattcaact   1320 

atattctagt cgtcggcgca aaagaggcag agtctggaaa ggtctctctg agggtaagag   1380 

acagggcaga cctatccaca gagagcattg ctgacgtcat tgcacgtttt aacgacgaag   1440 

ttgcgtcttt ccagtgattt ttaagtcgca tcatcttttt tttgtccaag acatctactg   1500 

cacaacccac attgtaattt ggtgaagtga ggtgaatgaa aaatcatgat attcttgttc   1560 

atgttgtcac atgtacatta actgccatga tgtatcaatt ctataagggc ctctttgatt   1620 

cgaaggattt tcatggggat tggaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaa      1677  
           
             28  
             483  
             PRT  
             Triticum aestivum  
           
            28 

Glu Asn Glu Phe Lys Val Glu Ile Ile Asn Glu Leu Pro Glu Asp Lys 
  1               5                  10                  15 

Thr Ile Thr Val Tyr Arg Cys Gly Pro Leu Val Asp Leu Cys Arg Gly 
             20                  25                  30 

Pro His Ile Pro Asn Thr Ser Phe Val Lys Ala Phe Ala Cys Leu Lys 
         35                  40                  45 

Ala Ser Ala Ser Tyr Trp Arg Gly Lys Ala Asp Arg Glu Ser Leu Gln 
     50                  55                  60 

Arg Val Tyr Gly Ile Ser Phe Pro Asp Ser Lys Arg Leu Lys Glu Tyr 
 65                  70                  75                  80 

Gln His Met Ile Glu Glu Ala Lys Lys Arg Asp His Arg Leu Leu Gly 
                 85                  90                  95 

Gln Ser Gln Lys Leu Phe Phe Phe His Pro Leu Ser Pro Gly Ser Cys 
            100                 105                 110 

Phe Phe Leu Pro Asn Gly Ala Ile Ile Tyr Asn Lys Leu Met Asp Phe 
        115                 120                 125 

Leu Arg Lys Glu Tyr Arg Glu Arg Gly Tyr Gln Glu Val Leu Ser Pro 
    130                 135                 140 

Asn Ile Tyr Asn Met Gln Leu Trp Glu Thr Ser Gly His Ala Ala Asn 
145                 150                 155                 160 

Tyr Lys Asp Asn Met Phe Val Phe Glu Ile Glu Lys Gln Glu Phe Gly 
                165                 170                 175 

Leu Lys Pro Met Asn Cys Pro Gly His Cys Leu Met Phe Gly His Glu 
            180                 185                 190 

Val Arg Ser Tyr Arg Glu Leu Pro Leu Arg Met Ala Asp Phe Gly Val 
        195                 200                 205 

Leu His Arg Asn Glu Leu Ser Gly Ala Leu Thr Gly Leu Thr Arg Val 
    210                 215                 220 

Arg Arg Phe Gln Gln Asp Asp Ala His Ile Phe Cys Met Glu Ser Gln 
225                 230                 235                 240 

Ile Lys Asp Glu Val Arg Ala Cys Leu Glu Phe Ile Asp Tyr Val Tyr 
                245                 250                 255 

Lys Ile Phe Gly Phe Glu Tyr Glu Leu Glu Leu Ser Thr Arg Pro Glu 
            260                 265                 270 

Lys Tyr Leu Gly Asp Ile Glu Thr Trp Asn Lys Ala Glu Gln Gln Leu 
        275                 280                 285 

Thr Glu Ala Leu Asn Glu Phe Gly Lys Pro Trp Lys Ile Asn Glu Ala 
    290                 295                 300 

Asp Gly Ala Phe Tyr Gly Pro Lys Ile Asp Ile Gly Val Phe Asp Ala 
305                 310                 315                 320 

Leu Lys Arg Lys Phe Gln Cys Ala Thr Leu Gln Leu Asp Phe Gln Leu 
                325                 330                 335 

Pro Leu Arg Phe Lys Leu Thr Tyr Ser Ala Glu Asp Glu Ala Lys Leu 
            340                 345                 350 

Glu Arg Pro Val Met Ile His Arg Ala Ile Leu Gly Ser Val Glu Arg 
        355                 360                 365 

Met Phe Ala Ile Leu Leu Glu His Tyr Asn Gly Lys Trp Pro Leu Trp 
    370                 375                 380 

Leu Ser Pro Arg Gln Ala Ile Val Cys Cys Val Ser Ala Asn Ser Leu 
385                 390                 395                 400 

Thr Tyr Ala Lys Glu Val His Ala Gln Ile Arg Ala Ala Gly Phe His 
                405                 410                 415 

Val Asp Ile Asp Met Thr Asp Arg Thr Ile Gln Lys Lys Val Arg Glu 
            420                 425                 430 

Ala Gln Leu Ala Gln Phe Asn Tyr Ile Leu Val Val Gly Ala Lys Glu 
        435                 440                 445 

Ala Glu Ser Gly Lys Val Ser Leu Arg Val Arg Asp Arg Ala Asp Leu 
    450                 455                 460 

Ser Thr Glu Ser Ile Ala Asp Val Ile Ala Arg Phe Asn Asp Glu Val 
465                 470                 475                 480 

Ala Ser Phe