Based on Mark's suggestion, I took a look at the EST files. Luckily there is no protein sequence contamination.<div><br></div><blockquote style="margin:0 0 0 40px;border:none;padding:0px"><div><div><font face="courier new, monospace">[dstandag@mason Transcriptome] grep -v '^>' Pdom.Trinity.Trimmomatic.fasta | grep -o . | sort | uniq -c</font></div>
</div><div><div><font face="courier new, monospace">79400764 A</font></div></div><div><div><font face="courier new, monospace">39834991 C</font></div></div><div><div><font face="courier new, monospace">40702954 G</font></div>
</div><div><div><font face="courier new, monospace">77980105 T</font></div></div><div><div><font face="courier new, monospace">[dstandag@mason Transcriptome] grep -v '^>' Pmet.Trinity.R.fasta | grep -o . | sort | uniq -c</font></div>
</div><div><div><font face="courier new, monospace">18294708 A</font></div></div><div><div><font face="courier new, monospace">9108213 C</font></div></div><div><div><font face="courier new, monospace">9449127 G</font></div>
</div><div><div><font face="courier new, monospace">17756470 T</font></div></div></blockquote><div class="gmail_extra"><br clear="all"><br>--<br>Daniel S. Standage<br>Ph.D. Candidate<br>Bioinformatics and Computational Biology Program<br>
Department of Genetics, Development, and Cell Biology<br>Iowa State University<br><br>
<br><br><div class="gmail_quote">On Thu, Nov 8, 2012 at 9:47 AM, Mark Yandell <span dir="ltr"><<a href="mailto:myandell@genetics.utah.edu" target="_blank">myandell@genetics.utah.edu</a>></span> wrote:<br><blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex">
Hi Daniel,<br>
<br>
is it possible you have some proteins in your EST files?<br>
<br>
'------------- EXCEPTION: Bio::Root::Exception -------------<br>
<div class="im">MSG: Sequence is a protein. Cannot revcom<br>
</div>STACK: Error::throw'<br>
<br>
<br>
<br>
Mark Yandell<br>
Professor of Human Genetics<br>
H.A. & Edna Benning Presidential Endowed Chair<br>
Eccles Institute of Human Genetics<br>
University of Utah<br>
15 North 2030 East, Room 2100<br>
Salt Lake City, UT 84112-5330<br>
ph:<a href="tel:801-587-7707" value="+18015877707">801-587-7707</a><br>
<br>
________________________________________<br>
From: <a href="mailto:maker-devel-bounces@yandell-lab.org">maker-devel-bounces@yandell-lab.org</a> [<a href="mailto:maker-devel-bounces@yandell-lab.org">maker-devel-bounces@yandell-lab.org</a>] on behalf of Daniel Standage [<a href="mailto:daniel.standage@gmail.com">daniel.standage@gmail.com</a>]<br>
Sent: Thursday, November 08, 2012 7:32 AM<br>
To: Carson Holt<br>
Cc: Maker Mailing List<br>
Subject: Re: [maker-devel] Maker issues<br>
<div><div class="h5"><br>
Scaling up to whole-genome annotation, things seem to be going well. However, there are some intermittent issues. I've seen a couple occurrences of the following error...<br>
<br>
#-------------------------------#<br>
Calling out to FastaSeq::convert at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/GI.pm line 1480.<br>
running est2genome search.<br>
#--------- command -------------#<br>
Widget::exonerate::est2genome:<br>
/N/hd01/dstandag/Mason/local/bin/exonerate -q /N/dc/scratch/dstandag/PdomGenomic/Annotation/output/maker.pdom.5.mason.maker.output/maker.pdom.5.mason_datastore/scaffold_23/theVoid.scaffold_23/comp58983_c0_seq101.for.716125-721460.0.fasta -t /N/dc/scratch/dstandag/PdomGenomic/Annotation/output/maker.pdom.5.mason.maker.output/maker.pdom.5.mason_datastore/scaffold_23/theVoid.scaffold_23/scaffold_23.716125-721460.0.fasta -Q dna -T dna --model est2genome --minintron 20 --maxintron 10000 --showcigar --percent 20 > /N/dc/scratch/dstandag/PdomGenomic/Annotation/output/maker.pdom.5.mason.maker.output/maker.pdom.5.mason_datastore/scaffold_23/theVoid.scaffold_23/scaffold_23.716125-721460.comp58983_c0_seq101.est_exonerate.0<br>
#-------------------------------#<br>
Calling out to FastaSeq::convert at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/GI.pm line 1480.<br>
couldn't close /N/dc/scratch/dstandag/PdomGenomic/Annotation/output/maker.pdom.5.mason.maker.output/maker.pdom.5.mason_datastore/scaffold_23/theVoid.scaffold_23/comp58983_c0_seq37.for.716125-723330.0.fasta at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/FastaFile.pm line 60.<br>
--> rank=NA, hostname=c4<br>
ERROR: Failed while polishig ESTs<br>
ERROR: Chunk failed at level:2, tier_type:2<br>
FAILED CONTIG:scaffold_23<br>
<br>
ERROR: Chunk failed at level:5, tier_type:0<br>
FAILED CONTIG:scaffold_23<br>
<br>
examining contents of the fasta file and run log<br>
Calling Datastore::MD5::mkdir at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm line 433.<br>
Calling uri_escape at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm line 433.<br>
Calling File::Path::mkpath at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm line 433.<br>
<br>
<br>
...as well as one occurrence of this error.<br>
<br>
#-------------------------------#<br>
Calling out to FastaSeq::convert at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/GI.pm line 1480.<br>
running est2genome search.<br>
#--------- command -------------#<br>
Widget::exonerate::est2genome:<br>
/N/hd01/dstandag/Mason/local/bin/exonerate -q /N/dc/scratch/dstandag/PdomGenomic/Annotation/output/maker.pdom.3.mason.maker.output/maker.pd<br>
om.3.mason_datastore/scaffold_7/theVoid.scaffold_7/comp59027_c1_seq93.for.1869077-1869882.0.fasta -t /N/dc/scratch/dstandag/PdomGenomic/Anno<br>
tation/output/maker.pdom.3.mason.maker.output/maker.pdom.3.mason_datastore/scaffold_7/theVoid.scaffold_7/scaffold_7.1869077-1869882.0.fasta<br>
-Q dna -T dna --model est2genome --minintron 20 --maxintron 10000 --showcigar --percent 20 > /N/dc/scratch/dstandag/PdomGenomic/Annotation/<br>
output/maker.pdom.3.mason.maker.output/maker.pdom.3.mason_datastore/scaffold_7/theVoid.scaffold_7/scaffold_7.1869077-1869882.comp59027_c1_se<br>
q93.est_exonerate.0<br>
#-------------------------------#<br>
<br>
------------- EXCEPTION: Bio::Root::Exception -------------<br>
MSG: Sequence is a protein. Cannot revcom<br>
STACK: Error::throw<br>
STACK: Bio::Root::Root::throw /N/u/dstandag/Mason/local/src/PerlLibs/lib/perl5/Bio/Root/Root.pm:368<br>
STACK: Bio::PrimarySeqI::revcom /N/u/dstandag/Mason/local/src/PerlLibs/lib/perl5/Bio/PrimarySeqI.pm:381<br>
STACK: Bio::LocatableSeq::revcom /N/u/dstandag/Mason/local/src/PerlLibs/lib/perl5/Bio/LocatableSeq.pm:577<br>
</div></div>STACK: exonerate::splice_info::needs_to_be_revcomped /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/exonerate/<a href="http://splice_info.pm:86" target="_blank">splice_info.pm:86</a><<a href="http://splice_info.pm:86" target="_blank">http://splice_info.pm:86</a>><br>
STACK: Widget::exonerate::est2genome::assemble /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Widget/exonerate/<a href="http://est2genome.pm:686" target="_blank">est2genome.pm:686</a><<a href="http://est2genome.pm:686" target="_blank">http://est2genome.pm:686</a>><br>
STACK: Widget::exonerate::est2genome::parse /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Widget/exonerate/<a href="http://est2genome.pm:961" target="_blank">est2genome.pm:961</a><<a href="http://est2genome.pm:961" target="_blank">http://est2genome.pm:961</a>><br>
STACK: polisher::exonerate::est::e_exonerate /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/polisher/exonerate/<a href="http://est.pm:82" target="_blank">est.pm:82</a><<a href="http://est.pm:82" target="_blank">http://est.pm:82</a>><br>
STACK: polisher::exonerate::est::polish /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/polisher/exonerate/<a href="http://est.pm:44" target="_blank">est.pm:44</a><<a href="http://est.pm:44" target="_blank">http://est.pm:44</a>><br>
<div><div class="h5">STACK: GI::to_polisher /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/GI.pm:1670<br>
STACK: GI::polish_exonerate /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/GI.pm:1517<br>
STACK: Process::MpiChunk::_go /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm:1663<br>
STACK: Process::MpiChunk::run /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm:335<br>
STACK: Process::MpiChunk::run_all /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm:351<br>
STACK: Process::MpiTiers::run_all /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiTiers.pm:286<br>
STACK: Process::MpiTiers::run_all /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiTiers.pm:286<br>
STACK: /N/u/dstandag/Mason/local/src/maker-dev/bin/maker:644<br>
-----------------------------------------------------------<br>
--> rank=NA, hostname=c4<br>
ERROR: Failed while polishig ESTs<br>
ERROR: Chunk failed at level:2, tier_type:2<br>
FAILED CONTIG:scaffold_7<br>
<br>
ERROR: Chunk failed at level:5, tier_type:0<br>
FAILED CONTIG:scaffold_7<br>
<br>
examining contents of the fasta file and run log<br>
Calling Datastore::MD5::mkdir at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm line 433.<br>
Calling uri_escape at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm line 433.<br>
Calling File::Path::mkpath at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm line 433.<br>
<br>
I'll let you know if I see anything else.<br>
<br>
<br>
--<br>
Daniel S. Standage<br>
Ph.D. Candidate<br>
Bioinformatics and Computational Biology Program<br>
Department of Genetics, Development, and Cell Biology<br>
Iowa State University<br>
<br>
<br>
<br>
</div></div><div class="im">On Wed, Nov 7, 2012 at 11:46 AM, Carson Holt <<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a><mailto:<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a>>> wrote:<br>
Thanks. Typo now fixed on my end too ;-)<br>
<br>
Thanks,<br>
Carson<br>
<br>
<br>
</div>From: Daniel Standage <<a href="mailto:daniel.standage@gmail.com">daniel.standage@gmail.com</a><mailto:<a href="mailto:daniel.standage@gmail.com">daniel.standage@gmail.com</a>>><br>
<div class="im">Date: Wednesday, 7 November, 2012 11:43 AM<br>
<br>
</div>To: Carson Holt <<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a><mailto:<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a>>><br>
Cc: Maker Mailing List <<a href="mailto:maker-devel@yandell-lab.org">maker-devel@yandell-lab.org</a><mailto:<a href="mailto:maker-devel@yandell-lab.org">maker-devel@yandell-lab.org</a>>><br>
<div class="im">Subject: Re: Maker issues<br>
<br>
Looked good for a while, but came across this error.<br>
<br>
total clusters:20 now processing 0<br>
flattening EST clusters<br>
doing tblastx of alt-ESTs<br>
Undefined subroutine &GI::loalize_file called at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/GI.pm line 2648.<br>
--> rank=NA, hostname=c4<br>
ERROR: Failed while doing tblastx of alt-ESTs<br>
ERROR: Chunk failed at level:4, tier_type:2<br>
FAILED CONTIG:scaffold_58<br>
<br>
ERROR: Chunk failed at level:5, tier_type:0<br>
FAILED CONTIG:scaffold_58<br>
<br>
examining contents of the fasta file and run log<br>
Calling Datastore::MD5::mkdir at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm line 433.<br>
Calling uri_escape at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm line 433.<br>
Calling File::Path::mkpath at /N/hd01/dstandag/Mason/local/src/maker-dev/bin/../lib/Process/MpiChunk.pm line 433.<br>
<br>
<br>
<br>
--Next Contig--<br>
<br>
It seems pretty clear that there is a typo in GI.pm. I changed loalize to localize and relaunched.<br>
<br>
<br>
--<br>
Daniel S. Standage<br>
Ph.D. Candidate<br>
Bioinformatics and Computational Biology Program<br>
Department of Genetics, Development, and Cell Biology<br>
Iowa State University<br>
<br>
<br>
<br>
</div><div class="im">On Wed, Nov 7, 2012 at 9:30 AM, Daniel Standage <<a href="mailto:daniel.standage@gmail.com">daniel.standage@gmail.com</a><mailto:<a href="mailto:daniel.standage@gmail.com">daniel.standage@gmail.com</a>>> wrote:<br>
Done.<br>
<br>
Test job has successfully cleared the preliminary Fasta indexing steps and is repeat masking. I'll let you know if there are any problems. Thanks!<br>
<br>
<br>
--<br>
Daniel S. Standage<br>
Ph.D. Candidate<br>
Bioinformatics and Computational Biology Program<br>
Department of Genetics, Development, and Cell Biology<br>
Iowa State University<br>
<br>
<br>
<br>
</div><div class="im">On Wed, Nov 7, 2012 at 9:00 AM, Carson Holt <<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a><mailto:<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a>>> wrote:<br>
1.006902 Bio::Root::Version /N/u/dstandag/Mason/local/src/PerlLibs/bioperl-live/Bio/Root/Version.pm<br>
<br>
One thing I noticed, in the debug output is that you are using Bioperl live (here --> /N/u/dstandag/Mason/local/src/PerlLibs/bioperl-live). It's fasta indexer is broken. I have an open bug I am trying to resolve with the Bioperl developers, but for now use the CPAN version of Bioperl.<br>
<br>
Thanks,<br>
Carson<br>
<br>
<br>
<br>
<br>
</div>From: Daniel Standage <<a href="mailto:daniel.standage@gmail.com">daniel.standage@gmail.com</a><mailto:<a href="mailto:daniel.standage@gmail.com">daniel.standage@gmail.com</a>>><br>
<div class="im">Date: Monday, 5 November, 2012 10:14 AM<br>
</div>To: Carson Holt <<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a><mailto:<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a>>><br>
Cc: Maker Mailing List <<a href="mailto:maker-devel@yandell-lab.org">maker-devel@yandell-lab.org</a><mailto:<a href="mailto:maker-devel@yandell-lab.org">maker-devel@yandell-lab.org</a>>><br>
<div class="im">Subject: Re: Maker issues<br>
<br>
Debug output attached (bzip2 compressed).<br>
<br>
<br>
--<br>
Daniel S. Standage<br>
Ph.D. Candidate<br>
Bioinformatics and Computational Biology Program<br>
Department of Genetics, Development, and Cell Biology<br>
Iowa State University<br>
<br>
<br>
<br>
</div><div class="im">On Mon, Nov 5, 2012 at 10:08 AM, Carson Holt <<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a><mailto:<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a>>> wrote:<br>
Thanks. Could you also run with the --debug flag set on the command line for a few minutes and send me that.<br>
<br>
--Carson<br>
<br>
<br>
</div>From: Daniel Standage <<a href="mailto:daniel.standage@gmail.com">daniel.standage@gmail.com</a><mailto:<a href="mailto:daniel.standage@gmail.com">daniel.standage@gmail.com</a>>><br>
<div class="im">Date: Monday, 5 November, 2012 10:05 AM<br>
</div>To: Carson Holt <<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a><mailto:<a href="mailto:carsonhh@gmail.com">carsonhh@gmail.com</a>>>, Maker Mailing List <<a href="mailto:maker-devel@yandell-lab.org">maker-devel@yandell-lab.org</a><mailto:<a href="mailto:maker-devel@yandell-lab.org">maker-devel@yandell-lab.org</a>>><br>
<div class="HOEnZb"><div class="h5">Subject: Maker issues<br>
<br>
Carson,<br>
<br>
I updated to the latest development version, made sure the TMP directory is on native disk space, and relaunched. I have attached the output of the job that failed in <5 minutes. It looks pretty similar to the errors I got the last time I used the dev version.<br>
<br>
--<br>
Daniel S. Standage<br>
Ph.D. Candidate<br>
Bioinformatics and Computational Biology Program<br>
Department of Genetics, Development, and Cell Biology<br>
Iowa State University<br>
<br>
<br>
<br>
<br>
<br>
</div></div></blockquote></div><br></div>