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Chalara fraxinea gene predictions, TGAC_Chalara_fraxinea_ass_s1v1_ann_v1.1

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swarbred edited this page Feb 18, 2013 · 2 revisions

Version 1.1 Chalara fraxinea gene predictions

ReadMe - TGAC_Chalara_fraxinea_ass_s1v1_ann_v1.1

Repeat Masking

RepeatModeler v1-0-7 was used to generate a species specific repeat library based on the TGAC s1v1 Chalara assembly. Interspersed repeats were identified using the Chalara fraxinea repeat library and RepeatMasker. Low complexity repeats were identified with RepeatMasker and tantan http://www.cbrc.jp/tantan/.

GFF files:

/Repeats/RM_int_repeats.gff

/Repeats/RM_loc_complex_repeats.gff

/Repeats/tantan_low_complex_repeats.gff

Evidence alignments:

Proteins

Protein sequences from 4 species Amorphotheca resinae v1.0, Botrytis cinerea v1.0, Oidiodendron maius Zn v1.0 and Sclerotinia sclerotiorum v1.0 (http://genome.jgi.doe.gov/pages/dynamicOrganismDownload.jsf?organism=leotiomycetes) were soft masked for low complexity (segmasker) and aligned to the soft masked TGAC s1v1 Chalara assembly with exonerate protein2genome, alignments were filtered at a minimum 50% identity and 50% coverage.

GFF files: Source/Amore1_GeneCatalog_proteins_20110719.aa.mask.exonerate.50id-50cov.gff Source/Botci1_GeneCatalog_proteins_20110903.aa.mask.exonerate.50id-50cov.gff Source/Oidma1_GeneCatalog_proteins_20110606.aa.mask.exonerate.50id-50cov.gff Source/Sclsc1_GeneCatalog_proteins_20110903.aa.mask.exonerate.50id-50cov.gff

RNA-Seq

The public RNA-Seq reads were aligned to the unmasked TGAC s1v1 Chalara assembly with Tophat v2.0.4 (--min-anchor-length 12 --max-multihits 20 --min-intron-length 50 --max-intron-length 10000 --min-coverage-intron 50 --max-coverage-intron 5000 --min-segment-intron 50 --max-segment-intron 10000).

reads: ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R1_001.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R1_002.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R1_003.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R1_004.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R1_005.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R1_006.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R2_001.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R2_002.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R2_003.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R2_004.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R2_005.fastq.gz ftp-oadb.tsl.ac.uk/mixed_material/ashwellthorpe_AT2/RNA_seq/lane5_NoIndex_L005_R2_006.fastq.gz

RNA-Seq data contains 23,096,021 PE reads of 76 bp of these 3,623,719 (16%) read pairs have at least 1 read of the pair mapping to the s1v1 Chalara assembly assembly subset (6,255,701 positions mapped 67% properly paired)

Splice junctions identified with Tophat were filtered requiring support from a minimum of 4 reads.

Junction file (unfiltered): Source/jasmbl_filtered.gbrowse.gff

Bigwig file of RNA-Seq read density: Source/accepted_hits.sort.bam.update.bw

RNA-Seq alignments were assembled with Cufflinks v2.0.2 (-I 10000 -A 0.15) into 11804 contigs.

  • stats_info bases 21858299
  • stats_info contigs 11804
  • stats_len max 17863
  • stats_len mean 1851.77
  • stats_len median 1492
  • stats_len min 86
  • stats_len mode 769
  • stats_len modeval 13
  • stats_len range 17778
  • stats_len stddev 1430.24

Longest ORFs were selected: Source/cufflinks_transcripts_cds.gff

The public Trinity Assembly of the same RNA-Seq data (AT2_trinity_version2) was softmasked (dustmasker) and aligned to the soft masked TGAC s1v1 Chalara assembly with exonerate est2genome and filtered at a minimum of 95% identity, 50% coverage.

GFF file: Source/AT2_trinity_version2.mask.fasta.exonerate.95id-50cov.gff

Augustus Training and genebuild

Augustus v2.5.5 was trained for Chalara_fraxinea using a subset of cufflinks assemblies. Briefly, blast searches against the 4 protein datasets (see above) were used to identify predicted cufflinks CDS features with support from cross species alignments, these were further filtered to remove features showing >80% sequence similarity within the selected subset and/or any genomic overlap. Augustus was trained based on the filtered set of 859 cufflinks models with 100 models reserved for testing. Based on this the ab initio predictions achieved sensitivity results of 0.96 nucleotide, 0.80 exon and 0.65 at the gene level.

Augustus gene models were predicted using the trained ab initio model with the 4 cross species protein alignments, RNA-Seq junctions, Cufflinks alignments, and Aligned Trinity contigs as evidence hints. RNA-Seq read density was provided as exon hints and repeat information as nonexonpart hints. Augustus gene models were filtered to remove redundant genes giving a total of 10252 predicted genes with 10405 predicted mRNAs.

GFF file:

Gene_predictions/TGAC_Chalara_fraxinea_ass_s1v1_ann_v1.1 /Chalara_fraxinea_ass_s1v1_ann_v1.1.gene.gff

Genemodels are assigned a uniq identifier of the following form

CHAFR746836.1.1_0000010.1 (uniq species code incorporating NCBI taxonomy id CHAFR746836, 1.1. annotation version, 0000010 gene identifier, .1 transcript identifier)

  • Gene Stats Total (bp) 23087543 Total Count 10252 Mean size (span) 2252
  • mRNA Stats Total (bp) 23462930 Total Count 10405 Mean size (span) 2254.97
  • Exon Stats (distinct) Total (bp) 20614305 Total Count 34735 Mean size 593.474
  • Intron Stats (distinct) Total (bp) 1849527 Total Count 22347 Mean size 82.764
  • CDS Exon Stats (distinct) Total (bp) 15661318 Total Count 32568 Mean size 480.881
  • 5UTR Exon Stats Total (bp) 2402584 Total Count 11954 Mean size 200.986
  • 3UTR Exon Stats Total (bp) 2598484 Total Count 10758 Mean size 241.54

cDNA transcripts stats Mean length 2000.53983661701 Median 1725 Min 204 Max 23327 Total Length 20815617

CDS transcript stats Mean length 1519.89082172033 Median 1272 Min 84 Max 22749 Total Length 15814464

Maker genebuild

An alternative genebuild was generated with maker-2.10 using the above evidence files and trained augustus model.

GFF file: Gene_predictions/TGAC_Chalara_fraxinea_ass_s1v1_ann_v1.1 /maker.genes.utr.gff

A Gbrowse instance is available at http://browser.tgac.bbsrc.ac.uk/cgi-bin/gb2/gbrowse/Chalara_fraxinea_ass_s1v1_ann_v1.1

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