dataesr/scientific-paragraphs-categorization huggingface dataset:
5 CNV per animal. The lowest number of CNVs was observed in the Red Jungle Fowl (Aviandiv) population with an average of 83. 8 CNV per animal, which was expected as an inbred Red Jungle Fowl animal was used as a reference in this study. The commercial broilers showed an average of 128. 8 CNVs per animal. These numbers are considerably higher than those reported by  Wang et al. (2012) , where the average was 40 CNVs per animal. Even when our analysis is restricted to the autosomes, as was done in the study of   respectively. The number of CNVRs in the chicken is comparable to that reported in human  [21]  and almost 3 times higher than in cattle  [22]. The 176-kb CNV linked to the late feathering locus  [14]  was detected in this dataset as CNVR 1508 on GGAZ between positions 9,971,185 and 10,140,048 with a size of 169 kb.
Gene content of chicken CNVR Within the 1,556 CNVRs, a total of 2,642 unique Ensembl peptides were identified based on chicken build 2. 1. To examine whether genes involved in specific pathways or biological processes are more prone to copy number variation, we performed a gene enrichment analysis for the genes located within the CNVRs. The chicken transcript ids were used as input into DAVID for a gene enrichment and ontology analysis  [26]. Terms showing significant enrichments were the GO terms "functional constituent of cytoskeleton", "nuclear binding", "cellular response to stress", and "macromolecule catabolic processes". The GO term "functional constituent of cytoskeleton" is mainly driven by the keratin superfamily. The avian keratin genes are over-represented when compared to mammals  [1].
aCGH data analysis and CNV calling Unamplified genomic DNA (1 μg) was labeled with Cy3 (test samples) or Cy5 (reference sample). The Agilent Oligonucleotide Array-based CGH for genomic DNA Analysis protocol (v4. 0:2006) was used for the labeling of the DNA, Hybridizations, washings, and scanning of the arrays. Self-self control hybridizations were performed by labeling the reference sample with Cy3 and Cy5. Fluorescence intensities ratios were extracted using Agilent Feature Extraction software (Agilent Technologies). Log2ratio profiles were then normalized using aCGH-Spline to remove dye biases and reduce experimental noise  [33]. CNV detection was performed using a modified version of CNVfinder  [32] , where we optimized empirically significance thresholds using replicate self-self hybridizations.
Acknowledgements This work was financially supported by  European Union  grant  FOOD-CT-2004-506416  (EADGENE),  BBSRC   Institute Strategic Programme Grants  at the  Pirbright Institute  and the  Roslin Institute, Hendrix Genetics, The Netherlands, Cobb-Vantress Inc, USA  and  The University of Delaware, College of Agriculture and Natural Resources Seed  grant  ANSC462123. We would like to thank  Carl J. Schmidt  and  Mary E. Delany  for their contribution of DNA from some of the lines and  Bert Dibbits  for the help in performing CNV validation.
Differential Scanning Calorimetry (DSC) DSC scans were made with a Mettler-Toledo (Viroflay, France) DSC calorimeter. The GNLs (10 mg) were dissolved in 100 μL of water and hermetically sealed in an aluminum pan. The temperature was increased at 0. 5 °C/min to 70 °C where it was held for 2 min. The temperature was then reduced to 10 °C and held at this temperature for 2 min. This heating-cooling cycle was repeated one more times before the sample was held isothermal at 10 °C for 20 min. In the case of oleyl derivative the heating-cooling cycle was done between -30 °C and to 70 °C. The data collected on the second cycle were analyzed. Integrating under the heat flow curve of the main peak afforded the enthalpy of the transition. Mettler Toledo STARe computer program was used to analyze the data.
subtilis are 51% identical with each other and 37% and 40% identical to the E. coli nagB gene product (Table  S1 ). Reizer et al  [3]  proposed that nagAB, together with the two domain transporter yflS, which they called nagP, were required for the metabolism of GlcNAc6P. (Confusingly, some databases refer to both the two-domain, yflF, and three-domain, ybfS, nagE homologues as nagP. To facilitate distinguishing the duplicated genes we have used the original nagAB, nagP and gamAP designations of Reizer et al  [3] , Figure  1 ). The third gene of the nagAB operon, yvoA, encodes a GntR family transcription factor. YvoA is homologous to the DasR protein, a pleiotropic regulator of use of GlcNAc and secondary metabolism in Streptomyces coelicolor  [8]. Bertram et al  [9]  showed, by transcriptome analysis, that nagA and nagP expression was increased in a yvoA mutant, which they proposed to call nagR.
mRNA Preparation, Northern Blots and Primer Extension Assays Bacteria were grown overnight in MOPS KGS medium with 0. 5% cas amino acids and then diluted into fresh medium with either Glc, GlcN GlcNAc or KGS as carbon source. Bacteria (25 ml) were harvested at an OD650 of about 0. 8 and frozen at 220uC. RNA was extracted by the glass beads and cold phenol method  [35]. RNAs were separated on 0. 8% agarose gels in 16 TBE and transferred overnight to HyBond N+ membranes by downward capillary transfer  [36]  in 56 SSC with 10 mM NaOH. The probes used were oligonucleotides of approximately 30-40 nt (Table  S2 ) labelled at their 59 end by polynucleotide kinase and c[ 32 P]ATP. Hybridization was carried out overnight using UltraHyb-oligo medium (Ambion) at 42uC. After washing, hybridization patterns were analysed by phosphoimagery and transcripts quantified with the ImageQuant program.
Acknowledgments We thank  Isabelle Martin-Verstraete ,  Josef Deutscher ,  Ciaran Condon ,  Patrick Stragier ,  Gerald Seidel ,  Didier Lereclus ,  Rainer Borriss  for gifts of strains and plasmids,  Peter Cherepanov  for information about pCP20,  Sylvain Durand  and  Laetitia Gilet  for advice on mRNA preparation and Northern analysis and  Annie Kolb  for critical comments on the manuscript.
EXPERIMENTAL PROCEDURES Crystallization and Data Collection-Recombinant SynCph2  (1) (2)  was produced as a histidine-tagged fusion with phycocyanobilin co-assembled in vivo. The expression and purification of SynCph2(1-2) was performed as previously described  (10). Crystals for the native datasets were grown using the hanging-drop vapor-diffusion method. The drops contained 1 l of 20 mg/ml of SynCph2(1-2)/P r -state in crys- tallization buffer (10 mM HEPES, 100 mM NaCl, pH 8. 0) and 1 l of reservoir solution (0. 1 M HEPES, pH 7. 0, 1. 0 M NH 4 COOH, 0. 1% (w/v) lysine, 0. 16% (w/v) arginine, 0. 05% (w/v) glutamate). Crystals were streak-seeded with a solution of crushed SynCph2(1-2) crystals. After pipetting the pre-irradiated (farred light, max ϭ 735 nm) protein solution under blue-safe light ( max ϭ 482 nm) conditions the plate was irradiated with farred light to ensure 100% occupancy of the P r state.
The crystals grew at 18 °C in the dark, were visualized under blue light, and frozen in reservoir buffer supplemented with 20% (v/v) glycerol. Selenomethionine-labeled SynCph2(1-2) crystals appeared with streak seeding using a reservoir solution of 0. 1 M HEPES, pH 6. 5, 0. 5 M NH 4 COOH and an initial protein concentration of 10 mg/ml. Native datasets were recorded at beamline ID14 -1 (European Synchrotron Radiation Facility (ESRF), Grenoble, France) and at beamline 14. 1 (Berliner Elektronenspeicherring-Gesellschaft fu ¨r Synchrotronstrahlung (BESSY-II), Berlin, Germany). The 3. 9-Å SAD dataset from the selenomethionine-labeled crystal was recorded at ID 14-4 (ESRF, Grenoble, France). Initial SAD phasing was performed with PHENIX (22) using the native 3. 20-Å dataset. Subsequent structural refinement with a 2.
60-Å dataset was executed with PHENIX and COOT  (23). Stereochemical restraints were configured for the PCB chromophore and its covalent attachment. Figures were created by using PYMOL 0. 99  (24). Single-crystal Resonance Raman Spectroscopy-Resonance Raman (RR) spectra were recorded offline at the Cryobench (European Synchrotron Radiation Facility (ESRF), Grenoble (France)) with a 785 nm near-IR laser using an inVia (Renishaw) Raman spectrometer (4 cm Ϫ1 spectral resolution)  (25). Resonance Raman spectra of the crystal (1 M MgOAc 2 as cryo-protectant) were collected under cryo-conditions (100 K) in the 200 -2000 cm Ϫ1 range using the SynchroScan method with a laser power of 300 milliwatts (50 milliwatts at the sample position) and 20-s exposure time. 30 -50 spectra were accumulated and baseline corrected to remove contributions from fluorescence and normalized to the 1642 cm Ϫ1 peak.
Although the QM fragment, consisting of the PCB chromophore, the Cys 129 side chain, and the pyrrole water that is bound to rings A, B, and C nitrogens, were described with the B3LYP density functional  (30) , the remaining protein atoms as well as solvation waters with the CHARMM22 force field  (31). The coupling between the QM and MM regions was described with the electrostatic embedding model combined with a charge shifted scheme  (32) , as implemented in the ChemShell software. The covalent bond cut at the QM/MM border on the Cys 129 was saturated with a hydrogen link atom. The optimization was performed using a limited memory quasi-Newton L-BFGS algorithm working with hybrid delocalized internal coordinates  (33). Production and Spectral Characterization of SynCph2(1-2) Variants-SynCph2(1-2) mutants were produced by QuikChange site-directed mutagenesis (Stratagene) of the pCDFDuet-1/SynCph2(1-2).
Acknowledgments-We thank the beamline staff of the  ESRF, Grenoble , and  BESSY-II, Berlin , for support,  Holger Steuber  for data collection,  Yann Geisselbrecht ,  Holger Steuber , and  Maik Veelders  for support in structure solution and refinement,  Petra Gnau  and  Ralf Po  ¨schke for technical support, and  Georgios Psakis  for discussions.
* This work was supported by  Deutsche Forschungsgemeinschaft  Grants  ES152/9-1  and  MR81/3-1  and the  LOEWE Center for Synthetic Microbiology  (to L. -O. E. ). □ S This article contains supplemental Figs. S1 and S2 and Tables  S1 andS2. The atomic coordinates and structure factors (code  4BWI ) have been deposited in the Protein Data Bank (http://wwpdb. org/).
RNA expression and transcriptome analysis Bioinformatics analysis of the datasets was as previously described and is publically available  [16]. Analysis of OPMDderegulated ECM pathways was performed with DAVID, functional annotation clustering tool  [17] , and statistical analysis of deregulated pathways was performed with Globaltest  [18]. PCOLCE expression values were obtained from a list of OPMD-deregulated top tables (p < 0. 05), comparing expression levels in VL muscles of OPMD and an age-matched control group  [16]. For validation studies reverse transcriptase quantitative-PCR (RT-qPCR) was performed on mRNA from either TA muscles of OPMD model and wild type control mice (A17. 1 and FVB mice respectively) or murine OPMD model myotube cultures (wild type IM2 and mutant Ala17). RNA extraction, cDNA synthesis and RT-qPCR analysis were performed as previously described  [16].
Collectively, our data from the The blot was incubated with anti-Pcolce antibody. Human PABPN1 is detected with an anti-FLAG antibody, the ACTA1 (MSA) protein is a marker for myotube differentiation and tubulin is used as a loading control. OPMD cell and animal model systems combined with that obtained from patient VL muscle biopsy samples, highlights a change in the sub-cellular distribution of PCOLCE in OPMD. In order to determine whether change in PCOLCE sub-cellular distribution is a general phenomenon of muscle wasting, PCOLCE sub-cellular localisation was studied in VL muscles of patients with DMD, myotonic dystrophy type 1 (DM1) and Facioscapulohumeral muscular dystrophy (FSHD). Histological analysis of muscle biopsies from all these conditions revealed widespread fibrosis and ECM thickening throughout muscle fibres (Figure  8A ).
Acknowledgements We thank  H. J. Ter Laak  ( Pathology Department, Nijmegen, Netherlands ) for providing patient muscle sections and  David C. Rubinsztein  for providing us with the A17. 1 OPMD mice. Our studies were supported by funds from the  European Commission  ( TRI-EX QLG2-CT-2001-01673  and  POLYALA LSHM-CT-2005-018675 ), the  Muscular Dystrophy Association  (Nr. 68015 ) and the  Association France for Myopathy  (Nr. 15 123 ).
Relaxation effect of Y-27632 Relaxation induced by Y-27632 [(R)-(+)-trans-N-(4-pyridyl)-4-(1-aminoethyl)-cyclohexanecarboxamide] (Tocris, France), a rho-kinase inhibitor, was tested on either the control rings or rings incubated in hemin following contraction induced by phenylephrine at 10 -6 M. Y-27632 was used at a concentration of 3 × 10 -7 M. Results were expressed as a percentage of the magnitude of contraction with 10 -6 M of phenylephrine. All the data were collected with a computerised dataacquisition system using Genie software (Adventech, USA). All the analyses were carried out using Origin 6 software (Microcal Software, Northampton, MA, USA).
Strain typing The phylogenetic group of the ESBL-producing E. coli was determined by a multiplex PCR assay  [18]. Isolates belonging to phylogenetic group B2 were screened with a previously established PCR-based method to identify the O25b subtype  [19]. Furthermore, multilocus sequence typing (MLST) using the scheme of the Institut Pasteur, Paris, France (www. pasteur. fr/mlst) was used to confirm that CTX-M-15-producing E. coli O25b belonged to the international clone ST131  [19]. Genetic relatedness of the ESBL-producing strains was studied by PFGE following extraction of genomic DNA and digestion with XbaI PFGE according to a standard protocol using a GenePath system (Bio-Rad). PFGE banding profiles were compared digitally using Fingerprint II software (Bio-Rad) and relatedness was calculated using the unweighted pair group method with arithmetic mean (UPGMA) algorithm with similarity of bands using the Dice similarity indices.
The NMD hypothesis also posits that introns should be inserted regularly along the gene. Indeed, a study of HGT genes in fungal genomes, mostly Ascomycetes, showed a correlation between intron densities in transgenes of bacterial origin and the recipient genomes  [34]. Here we studied an α-amylase gene, previously identified in a Basidiomycete, the white rot Phanerochaete chrysosporium  [35] , that was transferred from an Actinobacterium to Agaricomycotina. Alpha-amylases often form multigene families, and most Basidiomycetes already harbor at least one fungal-type α-amylase gene (Carbohydrate Active Enzymes database www. cazy. org [36] ). Basidiomycetes are ancestrally intron-rich  [7]. In this new gene of bacterial origin, we have identified intron gains and losses that occurred since the gene settled in the fungal genome and we estimated the rates of gains and losses, and some characteristics of the introns inserted.
Methods The sequence jgi|Phchr1|7087| from Phanerochaete chrysosporium, already reported to encode an animaltype α-amylase  [35]  was used as a query for BLASTP search in GenBank nr and GenBank Fungal Genomes (http://blast. ncbi. nlm. nih. gov), and BLASTP search implemented in the Mycocosm data base at the Joint Genome Institute (http://genome. jgi-psf. org/programs/fungi/, [37] ). The putative retrieved orthologs were then aligned using MAFFT  [38]  implemented in the Geneious software (Biomatters Ltd. ), and manually corrected for erroneous intronexon structures when necessary. Those errors were detected when large unique amino acid insertions or deletions were evidenced in the alignment. In these cases, when available and if necessary, expressed sequence tags (EST) were used to confirm intron positions and boundaries.
The query sequence contained a C-terminal carbohydrate binding module of the CBM20 family. A number, but not all retrieved sequences possessed a terminal CBM20 domain of variable length, always containing introns. Because it was not present in every sequence, the CBM20 was no longer considered and the alignment was truncated to the Cterminal end of the core protein. Intron positions were mapped onto the alignment according to the annotations of the genomes, mainly those deciphered at the JGI. From this protein alignment, after curation of the alignment with Gblocks  [39]  leaving 398 positions (83%) available, a gene tree was built using PhyML  [40] , at the www. phylogeny. fr web server  [41]. After testing various models with MEGA5  [42] , we used the WAG substitution matrix with a gamma distribution of substitution rate across sites (the shape parameter α was estimated from the data with four rate categories).
The robustness of the nodes was estimated by 100 bootstrap replicates. A few species were also investigated experimentally using polymerase chain reaction. DNA samples were supplied by the Hibbett Laboratory at Clark University or purchased from the Centraalbureau voor Schimmelcultures at the Institute of the Royal Netherlands Academy of Arts and Sciences. The primers and experimental conditions are given in Additional file 1: Table  S1. Only partial sequence data were obtained from the following species related to P. chrysosporium: Phlebia radiata FPL6140, P. albomellea CBS 275. 92, Grifola frondosa MO11 (accession numbers JX310736-JX310738). In order to infer the antiquity of the α-amylase gene transfer from a bacterium, and the times of intron insertions, we estimated the ages of nodes in a species tree.
A fungal species tree was established by compilation of recent literature, which included the species of interest for our study, but also Ascomycetes (  [43] [44] [45] [46] [47] [48] [49] [50] [51]  and especially  [52] ), and unpublished data kindly shared by D. S. Hibbett and by the Joint Genome Institute (Binder et al. in preparation) for solving uncertain relationships. We performed a Bayesian analysis with the BEAST program  [53]. An alignment was performed for 54 fungal species, using protein sequences of elongation factor 1-alpha, RNA polymerase II largest and second large subunits (EF1α, LSU1 and LSU2, Additional file 2: Table  S2 ) aligned separately using MAFFT  [38] , then concatenated. After curation for badly aligned regions with Gblocks  [39] , 1671 amino acid positions remained.
Because numerous gene duplications and gene losses occurred, we tried to reconcile the gene and species trees using Notung 2. 6  [62]. The program MALIN  [63]  infers the evolution of exon-intron structure in protein-coding orthologs. It could not be used, though, because orthology relationships among the genes could not be solved in most cases (see Results). Finally, the loss and gain events were mapped onto the species tree, not the gene tree. The average rates of intron gains and losses per million year and per branch were computed by considering that events occurred evenly along a branch. For example, if three losses occurred along a branch 12 Ma in length, the loss rate was 3/12 per Ma. Then, the rates for all branches present at a given time were summed and averaged.
Gene transfer from a bacterium We first performed TBLASTN and BLASTP searches against GenBank using the candidate α-amylase gene Phchr1|7087|. The best hits belonged to a few Basidiomycetes (Serpula lacrymans, Schizophyllum commune, Piriformospora indica) and then a lot of Bacteria, mainly Actinomycetales. No other fungus was found within the 100 first hits, except Moniliophthora perniciosa (Agaricales, Marasmiaceae), a truncated sequence which will be no longer considered here (MPER_11606), and a single Ascomycete species, Chaetomium globosum, already reported to harbor a similar α-amylase gene (CHGG_04966), but with a distinct bacterial origin  [35]. However, most fungal genome data have not been deposited yet to GenBank. Thus, we searched for genes similar to our P. chrysosporium query in the Mycocosm database at the JGI.
Our BLAST searches against all fungal database available to us (fungalgenomes. org) retrieved a total of 42 sequences with high similarity to the P. chrysosporium query (BLASTP expectvalue < 10 -109 in the Mycocosm database) from 24 species only, all Basidiomycetes. This confirmed the limited phylogenetic distribution of this gene among Fungi, and thus supported its likely bacterial origin. Figure  1  shows a tree of α-amylases of the GH13 family  [64]  from various prokaryotes and eukaryotes. This important enzyme group was divided in subfamilies  [58]. The tree shows that the genes we recovered in Basidiomycetes are grouped among Actinobacteria GH13_32 α-amylases, supporting an actinobacterial origin of the donor species. A very recent study supports this conclusion  [65]. The phylogenetic distribution of the recovered genes is limited to Agaricomycotina, suggests that the HGT event took place rather basally in Basidiomycetes, but after the split from Tremellomycetes, probably at the basal node of Agaricomycotina.
Intron richness, gains and losses As many as 480 introns that map at 64 intron positions were identified (Figure  2  and Additional file 4: Figure  S1 ). The number of introns per gene ranged from 8 (Ganoderma sp. Gansp1|123688|) to 22 (e. g. Stereum hirsutum Stehi1|95395|), with 13. 3 introns/gene on average, not counting the CBM20 extension. This high density considerably exceeds the average values for Basidiomycete genomes, which range from 3. 8 to 5. 7 introns/gene (data from the JGI). In addition, we found no correlation between the average genomic intron density and the intron density in the HGT Amy genes (not shown), in contrast to a previous study  [34]. This may be explained by the fact that all the species in our study fall within a relatively narrow range of genomic intron density, compared to the span of the study cited above, which included Ascomycetes, that are more intron-poor, and by the likely large within-genome variance.
below). Studying synteny among species (Mycocosm website) was no more helpful to uncover orthology relationships, because of quick loss of synteny, except in closely related species. Therefore, we attempted to reconcile the gene tree with the known phylogeny using the Notung software  [62]. We obtained a complex history, with 19 duplications and 56 gene losses (default parameters, with rearrangement option and rooting with Stehi1|78757|, Additional file 6: Figure  S3 ). Moreover, some major branches were marked as weak by the program. This may be due to the low support values at a majority of nodes (Additional file 5: Figure  S2 ). Therefore, we mapped the intron gains and losses on a species tree (Figure  3 ) from the data of Figure  2 , in a weighted parsimony framework. With this method, the possible parallel gains were limited to positions 2, 4, 21 and 24.
Acknowledgments Most of the sequence data were produced by the  US Department of Energy Joint Genome Institute  (http://www. jgi. doe. gov/) in collaboration with the user community. We are grateful to the  JGI  team and  Igor Grigoriev  for sharing unpublished data. We warmly thank  David Hibbett  and  Ricardo Garcia-Sandoval  at  Clark University  for their help, especially by giving DNA samples and sharing unpublished results. We thank  Dan Cullen  for advice regarding P. chrysosporium. We thank  Julien Fumey  for computer simulations. This work was funded by regular funding of the CNRS. SJ was supported by the grant No. 2/0148/11  from the Slovak grant agency  VEGA.
Size Exclusion Chromatography Coupled to Multiangle Laser Light Scattering (SEC-MALLS)-For SEC-MALLS analysis, MARS (0. 02 ml at 1 mg/ml) was loaded on a 300 ϫ 4. 6-mm BioSep-SEC-s4000 column (Phenomenex) equilibrated in 20 mM Hepes, pH 6. 8, 200 mM NaCl, 2% glycerol, and 2 mM DTT and developed at room temperature on a Prominence HPLC system (Shimadzu) at a flow rate of 0. 2 ml/min. Detection was performed using a three-detector static light scattering apparatus (MiniDAWN TREOS, Wyatt Technology, equipped with a quasielastic light scattering module) and a refractometer (Optilab rEX, Wyatt Technology). Molecular weight calculations were performed with the ASTRA V software (Wyatt Technology) using a dn/dc (Refractive Index Increment) value of 0. 183 ml/g. Dynamic Light Scattering-Hydrodynamic radius (Rh) measurements were made at 20 °C with a DynaPro DLS instrument (Protein Solutions Inc.