substr outside of string in PhatHits_utils.pm

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substr outside of string in PhatHits_utils.pm

Ole Kristian Tørresen-2
Dear all,
I'm having an issue with MAKER which I'm unable to wrap my head around. Hopefully the issue is easily identifiable and resolvable for someone with more insight than me. Please find the log output attached below. I cannot find any more information than this in any logs. Many scaffolds do complete fine, but some of the longest ones have issues.

Thank you.

Sincerely,
Ole K. Tørresen

Error message:

#--------- command -------------#
Widget::augustus:
/projects/cees/bin/augustus/augustus-3.2.3/bin/augustus  --strand=backward --species=gadMor2_code_braker2 --UTR=off --hintsfile=/tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_brak
er2.auto_annotator.xdef.augustus --extrinsicCfgFile=/projects/cees/bin/augustus/augustus-3.2.3/config/extrinsic/extrinsic.MPE.cfg --AUGUSTUS_CONFIG_PATH=/projects/cees/bin/augustus/augustus-3.2
.3/config /tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_braker2.auto_annotator.augustus.fasta > /tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_braker2.auto_annotato
r.augustus
#-------------------------------#
deleted:0 genes
begin called get_best_alt_splices1
 ...processing 0 of 2
 ...processing 1 of 2
end called get_best_alt_splices1
 ...processing 0 of 20
 ...processing 1 of 20
 ...processing 2 of 20
 ...processing 3 of 20
 ...processing 4 of 20
 ...processing 5 of 20
 ...processing 6 of 20
 ...processing 7 of 20
 ...processing 8 of 20
 ...processing 9 of 20
 ...processing 10 of 20
 ...processing 11 of 20
 ...processing 12 of 20
 ...processing 13 of 20
 ...processing 14 of 20
 ...processing 15 of 20
 ...processing 16 of 20
 ...processing 17 of 20
 ...processing 18 of 20
 ...processing 19 of 20
substr outside of string at /projects/cees/bin/maker/maker-3.1.1/bin/../lib/PhatHit_utils.pm line 850.
--> rank=NA, hostname=compute-31-18.local
ERROR: Failed while annotating transcripts
ERROR: Chunk failed at level:1, tier_type:4
FAILED CONTIG:GmG20150304_scaffold_8692

ERROR: Chunk failed at level:6, tier_type:0
FAILED CONTIG:GmG20150304_scaffold_8692

examining contents of the fasta file and run log
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Re: substr outside of string in PhatHits_utils.pm

Carson Holt-2
My first guess is that if you are using gff3 files as input to anything, then there may be an  issue with your GFF3 file. My second suggestion is to try MAKER 3.02.02 to see if it has the same issue.

—Carson


> On Nov 9, 2017, at 2:44 AM, Ole Kristian Tørresen <[hidden email]> wrote:
>
> Dear all,
> I'm having an issue with MAKER which I'm unable to wrap my head around. Hopefully the issue is easily identifiable and resolvable for someone with more insight than me. Please find the log output attached below. I cannot find any more information than this in any logs. Many scaffolds do complete fine, but some of the longest ones have issues.
>
> Thank you.
>
> Sincerely,
> Ole K. Tørresen
>
> Error message:
>
> #--------- command -------------#
> Widget::augustus:
> /projects/cees/bin/augustus/augustus-3.2.3/bin/augustus  --strand=backward --species=gadMor2_code_braker2 --UTR=off --hintsfile=/tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_brak
> er2.auto_annotator.xdef.augustus --extrinsicCfgFile=/projects/cees/bin/augustus/augustus-3.2.3/config/extrinsic/extrinsic.MPE.cfg --AUGUSTUS_CONFIG_PATH=/projects/cees/bin/augustus/augustus-3.2
> .3/config /tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_braker2.auto_annotator.augustus.fasta > /tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_braker2.auto_annotato
> r.augustus
> #-------------------------------#
> deleted:0 genes
> begin called get_best_alt_splices1
> ...processing 0 of 2
> ...processing 1 of 2
> end called get_best_alt_splices1
> ...processing 0 of 20
> ...processing 1 of 20
> ...processing 2 of 20
> ...processing 3 of 20
> ...processing 4 of 20
> ...processing 5 of 20
> ...processing 6 of 20
> ...processing 7 of 20
> ...processing 8 of 20
> ...processing 9 of 20
> ...processing 10 of 20
> ...processing 11 of 20
> ...processing 12 of 20
> ...processing 13 of 20
> ...processing 14 of 20
> ...processing 15 of 20
> ...processing 16 of 20
> ...processing 17 of 20
> ...processing 18 of 20
> ...processing 19 of 20
> substr outside of string at /projects/cees/bin/maker/maker-3.1.1/bin/../lib/PhatHit_utils.pm line 850.
> --> rank=NA, hostname=compute-31-18.local
> ERROR: Failed while annotating transcripts
> ERROR: Chunk failed at level:1, tier_type:4
> FAILED CONTIG:GmG20150304_scaffold_8692
>
> ERROR: Chunk failed at level:6, tier_type:0
> FAILED CONTIG:GmG20150304_scaffold_8692
>
> examining contents of the fasta file and run log
> _______________________________________________
> maker-devel mailing list
> [hidden email]
> http://box290.bluehost.com/mailman/listinfo/maker-devel_yandell-lab.org


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Re: substr outside of string in PhatHits_utils.pm

Ole Kristian Tørresen-2
Thank you Carson.

After a bit of struggling, I can confirm that the same error occurs in MAKER 3.01.2 (I guess you meant that version, couldn’t find 3.02.02).

I am providing a GFF to est_gff, with match and match_part entries. For at least one of the scaffolds, the last coordinate (column 5) is the same number as the length of the scaffold. That should be allowed by the GFF3 standard, right?

How can I troubleshoot this? The error message is not so informative. It seems that PhatHit_utils.pm tries to find a stop codon. Snipped from that file, lines 849-850:
#fix stop codon by walking downstream
my $has_stop = $tM->is_ter_codon(substr($transcript_seq, $end-1-3, 3));

The GFF I am using was the output of Mikado (https://www.biorxiv.org/content/early/2017/11/09/216994), which is GFF3, and then processed a bit to make it suitable for MAKER. First converted to GTF by 'mikado util convert mikado.loci.gff3 mikado.loci.gtf'

Then I selected only mRNA and exon entries, and changed mRNA to transcript to make it look like cufflinks output (and set a dummy score):
grep -P "\tmRNA\t|\texon\t" mikado.loci.gtf |sed "s/mRNA/transcript/g" |awk -F "\t" '{$9=$9"cov \"10.0\";"; OFS="\t"; print $1, $2, $3, $4, $5, $6, $7, $8, $9}' > mikado.loci.score.gtf

Before converting with cufflinks2gff3:
cufflinks2gff3 mikado.loci.score.gtf > ests.score.gff3

Thank you.

Ole


> On 09 Nov 2017, at 17:28, Carson Holt <[hidden email]> wrote:
>
> My first guess is that if you are using gff3 files as input to anything, then there may be an  issue with your GFF3 file. My second suggestion is to try MAKER 3.02.02 to see if it has the same issue.
>
> —Carson
>
>
>> On Nov 9, 2017, at 2:44 AM, Ole Kristian Tørresen <[hidden email]> wrote:
>>
>> Dear all,
>> I'm having an issue with MAKER which I'm unable to wrap my head around. Hopefully the issue is easily identifiable and resolvable for someone with more insight than me. Please find the log output attached below. I cannot find any more information than this in any logs. Many scaffolds do complete fine, but some of the longest ones have issues.
>>
>> Thank you.
>>
>> Sincerely,
>> Ole K. Tørresen
>>
>> Error message:
>>
>> #--------- command -------------#
>> Widget::augustus:
>> /projects/cees/bin/augustus/augustus-3.2.3/bin/augustus  --strand=backward --species=gadMor2_code_braker2 --UTR=off --hintsfile=/tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_brak
>> er2.auto_annotator.xdef.augustus --extrinsicCfgFile=/projects/cees/bin/augustus/augustus-3.2.3/config/extrinsic/extrinsic.MPE.cfg --AUGUSTUS_CONFIG_PATH=/projects/cees/bin/augustus/augustus-3.2
>> .3/config /tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_braker2.auto_annotator.augustus.fasta > /tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_braker2.auto_annotato
>> r.augustus
>> #-------------------------------#
>> deleted:0 genes
>> begin called get_best_alt_splices1
>> ...processing 0 of 2
>> ...processing 1 of 2
>> end called get_best_alt_splices1
>> ...processing 0 of 20
>> ...processing 1 of 20
>> ...processing 2 of 20
>> ...processing 3 of 20
>> ...processing 4 of 20
>> ...processing 5 of 20
>> ...processing 6 of 20
>> ...processing 7 of 20
>> ...processing 8 of 20
>> ...processing 9 of 20
>> ...processing 10 of 20
>> ...processing 11 of 20
>> ...processing 12 of 20
>> ...processing 13 of 20
>> ...processing 14 of 20
>> ...processing 15 of 20
>> ...processing 16 of 20
>> ...processing 17 of 20
>> ...processing 18 of 20
>> ...processing 19 of 20
>> substr outside of string at /projects/cees/bin/maker/maker-3.1.1/bin/../lib/PhatHit_utils.pm line 850.
>> --> rank=NA, hostname=compute-31-18.local
>> ERROR: Failed while annotating transcripts
>> ERROR: Chunk failed at level:1, tier_type:4
>> FAILED CONTIG:GmG20150304_scaffold_8692
>>
>> ERROR: Chunk failed at level:6, tier_type:0
>> FAILED CONTIG:GmG20150304_scaffold_8692
>>
>> examining contents of the fasta file and run log
>> _______________________________________________
>> maker-devel mailing list
>> [hidden email]
>> http://box290.bluehost.com/mailman/listinfo/maker-devel_yandell-lab.org
>

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Re: substr outside of string in PhatHits_utils.pm

Jacques Dainat-3
Dear Carson,

I got exactly the same problem as Ole Kristian Torresen.
substr outside of string at /projects/cees/bin/maker/maker-3.01.1/bin/../lib/PhatHit_utils.pm line 850.
I also tried with the version 3.00.0 and got the same problem.

My run:
I was using gff3 alignements as gff ESTs, proteins and activating repeat masking and trna.

The code line in PhatHit_utils.pm is the following:
      my $transcript_seq  = maker::auto_annotator::get_transcript_seq($hit, $seq); # few line before
      …..
  #fix stop codon by walking downstream 
  my $has_stop = $tM->is_ter_codon(substr($transcript_seq, $end-1-3, 3));   # <= line 850

So, I relaunched using the —debug option but didn’t find anything useful.

Consequently I modified the  code to display the $transcript_seq object.
I joined the log were you can see the printed object from line 10955. 
I also printed the size of the sequence (length($transcript_seq)) and $end.
We can see that it crashes because the sequence is 1369 bp long and we try to extract a stop codon from position 1372 (1376-1-3). 


Printing the object I saw that the bug looks to occur when handling the protein evidence Q8J0D9.1.
I checked if relaunching Maker using only this protein. But everything went fine.
Could it be the EST evidence that interfers how MAKER is handling the Q8J0D9.1 protein evidence ?

I hope you could help us.

Best regards,


Jacques Dainat, PhD
NBIS (National Bioinformatics Infrastructure Sweden)
Genome Annotation Service

Address:
Uppsala University, Biomedicinska Centrum
Department of Medical Biochemistry Microbiology, Genomics
Husargatan 3, box 582
S-75123 Uppsala Sweden
Phone: +46 18 471 46 25

On 21 Nov 2017, at 14:57, Ole Kristian Tørresen <[hidden email]> wrote:

Thank you Carson.

After a bit of struggling, I can confirm that the same error occurs in MAKER 3.01.2 (I guess you meant that version, couldn’t find 3.02.02).

I am providing a GFF to est_gff, with match and match_part entries. For at least one of the scaffolds, the last coordinate (column 5) is the same number as the length of the scaffold. That should be allowed by the GFF3 standard, right?

How can I troubleshoot this? The error message is not so informative. It seems that PhatHit_utils.pm tries to find a stop codon. Snipped from that file, lines 849-850:
#fix stop codon by walking downstream
my $has_stop = $tM->is_ter_codon(substr($transcript_seq, $end-1-3, 3));

The GFF I am using was the output of Mikado (https://www.biorxiv.org/content/early/2017/11/09/216994), which is GFF3, and then processed a bit to make it suitable for MAKER. First converted to GTF by 'mikado util convert mikado.loci.gff3 mikado.loci.gtf'

Then I selected only mRNA and exon entries, and changed mRNA to transcript to make it look like cufflinks output (and set a dummy score):
grep -P "\tmRNA\t|\texon\t" mikado.loci.gtf |sed "s/mRNA/transcript/g" |awk -F "\t" '{$9=$9"cov \"10.0\";"; OFS="\t"; print $1, $2, $3, $4, $5, $6, $7, $8, $9}' > mikado.loci.score.gtf

Before converting with cufflinks2gff3:
cufflinks2gff3 mikado.loci.score.gtf > ests.score.gff3

Thank you.

Ole


On 09 Nov 2017, at 17:28, Carson Holt <[hidden email]> wrote:

My first guess is that if you are using gff3 files as input to anything, then there may be an  issue with your GFF3 file. My second suggestion is to try MAKER 3.02.02 to see if it has the same issue.

—Carson


On Nov 9, 2017, at 2:44 AM, Ole Kristian Tørresen <[hidden email]> wrote:

Dear all,
I'm having an issue with MAKER which I'm unable to wrap my head around. Hopefully the issue is easily identifiable and resolvable for someone with more insight than me. Please find the log output attached below. I cannot find any more information than this in any logs. Many scaffolds do complete fine, but some of the longest ones have issues.

Thank you.

Sincerely,
Ole K. Tørresen

Error message:

#--------- command -------------#
Widget::augustus:
/projects/cees/bin/augustus/augustus-3.2.3/bin/augustus  --strand=backward --species=gadMor2_code_braker2 --UTR=off --hintsfile=/tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_brak
er2.auto_annotator.xdef.augustus --extrinsicCfgFile=/projects/cees/bin/augustus/augustus-3.2.3/config/extrinsic/extrinsic.MPE.cfg --AUGUSTUS_CONFIG_PATH=/projects/cees/bin/augustus/augustus-3.2
.3/config /tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_braker2.auto_annotator.augustus.fasta > /tmp/18899594.d/maker_m8xwVp/0/34_2.22239-28465.gadMor2_code_braker2.auto_annotato
r.augustus
#-------------------------------#
deleted:0 genes
begin called get_best_alt_splices1
...processing 0 of 2
...processing 1 of 2
end called get_best_alt_splices1
...processing 0 of 20
...processing 1 of 20
...processing 2 of 20
...processing 3 of 20
...processing 4 of 20
...processing 5 of 20
...processing 6 of 20
...processing 7 of 20
...processing 8 of 20
...processing 9 of 20
...processing 10 of 20
...processing 11 of 20
...processing 12 of 20
...processing 13 of 20
...processing 14 of 20
...processing 15 of 20
...processing 16 of 20
...processing 17 of 20
...processing 18 of 20
...processing 19 of 20
substr outside of string at /projects/cees/bin/maker/maker-3.1.1/bin/../lib/PhatHit_utils.pm line 850.
--> rank=NA, hostname=compute-31-18.local
ERROR: Failed while annotating transcripts
ERROR: Chunk failed at level:1, tier_type:4
FAILED CONTIG:GmG20150304_scaffold_8692

ERROR: Chunk failed at level:6, tier_type:0
FAILED CONTIG:GmG20150304_scaffold_8692

examining contents of the fasta file and run log
_______________________________________________
maker-devel mailing list
[hidden email]
http://box290.bluehost.com/mailman/listinfo/maker-devel_yandell-lab.org


_______________________________________________
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[hidden email]
http://box290.bluehost.com/mailman/listinfo/maker-devel_yandell-lab.org


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[hidden email]
http://box290.bluehost.com/mailman/listinfo/maker-devel_yandell-lab.org

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