failed to transcriptom indexing with tophat2

Question

[zizigolu]

Sorry guys,

Please some one help me to know why I face with error whenever I'm trying this syntax?

[izadi@lbox161 bowtie2-2.2.5]$ $TOP/tophat2 -G yeast_SGD_anno.gtf --transcriptome-index=transcriptome Saccharomyces_cerevisiae.R64-1-1.pep.all.bt2idx

[2015-09-03 14:36:44] Building transcriptome files with TopHat v2.1.0
-----------------------------------------------
[2015-09-03 14:36:44] Checking for Bowtie
          Bowtie version:     2.2.5.0
[2015-09-03 14:36:44] Checking for Bowtie index files (genome)..
[2015-09-03 14:36:44] Checking for reference FASTA file
[2015-09-03 14:36:44] Building transcriptome data files transcriptome/yeast_SGD_anno
[2015-09-03 14:36:44] Building Bowtie index from yeast_SGD_anno.fa
    [FAILED]
Error: Couldn't build bowtie index with err = 1
[izadi@lbox161 bowtie2-2.2.5]$

[Devon Ryan]

You'll have to go through the logs, manually execute the last command that tophat2 issued, and look at the error message that that gives.

[zizigolu]

thanks Devon,

in logs:

[2015-09-03 10:56:10] Building transcriptome files with TopHat v2.1.0
-----------------------------------------------
[2015-09-03 10:56:10] Checking for Bowtie
  Bowtie 2 not found, checking for older version..
Error: Bowtie not found on this system.

and

./tophat -G yeast_SGD_anno.gtf --transcriptome-index=transcriptome orf_coding.fasta

[Devon Ryan]

Wrong log, there's one that lists the exact commands being run.

[zizigolu]

sorry there are:

bowtie_inspect_recons.log in which:

bowtie-inspect: bt2_inspect.cpp:234: void print_ref_sequences(std::ostream&, bool, const EList<std::basic_string>char, std::char_traits<char>, std::allocator<char> >, 128>&, const TIndexOffU*, const std::string&): Assertion `0' failed.

tophat.log in which:

[2015-09-03 14:36:44] Building transcriptome files with TopHat v2.1.0
-----------------------------------------------
[2015-09-03 14:36:44] Checking for Bowtie
          Bowtie version:     2.2.5.0
[2015-09-03 14:36:44] Checking for Bowtie index files (genome)..
[2015-09-03 14:36:44] Checking for reference FASTA file
[2015-09-03 14:36:44] Building transcriptome data files transcriptome/yeast_SGD_anno
[2015-09-03 14:36:44] Building Bowtie index from yeast_SGD_anno.fa
    [FAILED]
Error: Couldn't build bowtie index with err = 1

run.log in which:

/usr/data/nfs6/izadi/test/tophat-2.1.0.Linux_x86_64/tophat -G yeast_SGD_anno.gtf --transcriptome-index=transcriptome Saccharomyces_cerevisiae.R64-1-1.pep.all.bt2idx
/usr/data/nfs6/izadi/test/tophat-2.1.0.Linux_x86_64/gtf_to_fasta --min-anchor 8 --splice-mismatches 0 --min-report-intron 50 --max-report-intron 500000 --min-isoform-fraction 0.15 --output-dir ./tophat_out/ --max-multihits 20 --max-seg-multihits 40 --segment-length 25 --segment-mismatches 2 --min-closure-exon 100 --min-closure-intron 50 --max-closure-intron 5000 --min-coverage-intron 50 --max-coverage-intron 20000 --min-segment-intron 50 --max-segment-intron 500000 --read-mismatches 2 --read-gap-length 2 --read-edit-dist 2 --read-realign-edit-dist 3 --max-insertion-length 3 --max-deletion-length 3 -z gzip --gtf-annotations transcriptome/yeast_SGD_anno.gff --no-closure-search --no-coverage-search --no-microexon-search transcriptome/yeast_SGD_anno.gff Saccharomyces_cerevisiae.R64-1-1.pep.all.bt2idx.fa transcriptome/yeast_SGD_anno.fa > ./tophat_out/logs/g2f.out
./bowtie2-build transcriptome/yeast_SGD_anno.fa transcriptome/yeast_SGD_anno

and the file named g2f.err that I could not open

[Devon Ryan]

If transcriptome/yeast_SGD_anno.fa still exists, then have a look at it. Presumably it has some sort of issue.

[zizigolu]

yeah Devon it is in transcriptome file in bowtie2 folder but i could not open that