[ensembl-dev] Codon called wrong in VEP when using custom build cache
Heidi Viitaniemi
hmviit at utu.fi
Wed Mar 27 12:05:51 GMT 2013
Hi Will,
Just as a follow up, the problem was the old version of the bioperl
which is in the INSTALL.pl.
Re-installing VEP with a link to pointing to a newer version of bioperl
did result to the same output with my custom cache asyou got with your run.
Thanks for all your help,
Heidi
27.3.2013 8:53, Heidi Viitaniemi kirjoitti:
> Hi Will,
>
> And thank you very much for looking into this.
>
> The reason why I want to build my own cahce from gtf and fasta of the
> Gasterosteus_aculeatus is that on groupXIX the two last (of the three
> ) supercontigs are flipped in the Ensembl genome (Ross&Peichel 2008).
> I also first thougt that reverse complementing the fasta and gtf for
> groupXIX was the problem but ENSGACG00000003129 is located on a
> reagion that I didn't touch in the gtf or the fasta.
>
> I'll check the versions of Bioperl in our server and try using only
> the fasta and gtf for ENSGACG00000003129.
>
> Thanks,
> Heidi
>
>
> 26.3.2013 16:04, Will McLaren kirjoitti:
>> Hi Heidi,
>>
>> Thanks for your patience, I've had a chance to look at this now.
>>
>> If I build a cache file from the following files:
>>
>> ftp://ftp.ensembl.org/pub/release-70/gtf/gasterosteus_aculeatus/Gasterosteus_aculeatus.BROADS1.70.gtf.gz
>>
>> and
>>
>> ftp://ftp.ensembl.org/pub/release-70/fasta/gasterosteus_aculeatus/dna/Gasterosteus_aculeatus.BROADS1.70.dna.toplevel.fa.gz
>>
>> I get (I think!) the correct output from the VEP:
>>
>> perl gtf2vep.pl <http://gtf2vep.pl> -i
>> Gasterosteus_aculeatus.BROADS1.70.gtf.gz -fasta
>> Gasterosteus_aculeatus.BROADS1.70.dna.toplevel.fa -species
>> gasterosteus_aculeatus -dir test/ -db 70
>> perl variant_effect_predictor.pl <http://variant_effect_predictor.pl>
>> -i gastero_in.txt -species gasterosteus_aculeatus -force -off -dir
>> test/ -db 70
>> grep -v # variant_effect_output.txt
>>
>> groupXIX_2822477_C/T groupXIX:2822477 T ENSGACG00000003129
>> ENSGACT00000004109 Transcript missense_variant 67
>> 49 17 A/T Gcg/Acg -
>> groupXIX_2822500_T/C groupXIX:2822500 C ENSGACG00000003129
>> ENSGACT00000004109 Transcript missense_variant 44
>> 26 9 D/G gAc/gGc -
>> groupXIX_2822523_C/T groupXIX:2822523 T ENSGACG00000003129
>> ENSGACT00000004109 Transcript initiator_codon_variant
>> 21 3 1 M/I atG/atA -
>> groupXIX_2822541_T/A groupXIX:2822541 A ENSGACG00000003129
>> ENSGACT00000004109 Transcript 5_prime_UTR_variant
>> 3 - - - - -
>>
>> This works the same if I use the version 67 files as it appears you have.
>>
>> So I suspect there is something different about your FASTA file - you
>> could check that the sequence of the groupXIX file matches that in
>> the file I link to above (do an md5sum or some such thing).
>>
>> It is also possible that an issue with older versions of BioPerl is
>> to blame - there was a known bug in the way BioPerl indexes large
>> FASTA file. Normally for Ensembl we recommend using BioPerl 1.2.3
>> (which contains the bug), but VEP works fine with the latest version.
>> I'd try updating your BioPerl install to the latest version, remove
>> the *.fa.index file that is generated next to your .fa file, and try
>> re-running gtf2vep.pl <http://gtf2vep.pl>
>>
>> Beyond this it's hard to say what's happening without seeing the
>> contents of your GTF and FASTA files. If the problem persists,
>> perhaps you could just pull out the lines in the GTF for
>> ENSGACT00000004109 and the sequence for groupXIX and if that still
>> gives you the same problem, send them to me so I can debug.
>>
>> Hope this helps!
>>
>> Will
>>
>>
>> On 19 March 2013 12:07, Heidi Viitaniemi <hmviit at utu.fi
>> <mailto:hmviit at utu.fi>> wrote:
>>
>> Hi Will,
>>
>> And thank you for your response. I'll wait for the solution. I
>> like the idea that you can incorporate your own data to run VEP.
>>
>> Thanks,
>> Heidi Viitaniemi
>>
>>
>>
>> 19.3.2013 13:42, Will McLaren kirjoitti:
>>> Hello Heidi,
>>>
>>> Thanks for finding this - the causes of this bug are I believe
>>> somewhat complex so may take a while to get to the bottom of it.
>>>
>>> Just wanted to let you know that your mail is not being ignored!
>>>
>>> Regards
>>>
>>> Will McLaren
>>> Ensembl Variation
>>>
>>>
>>> On 18 March 2013 13:48, Heidi Viitaniemi <hmviit at utu.fi
>>> <mailto:hmviit at utu.fi>> wrote:
>>>
>>> Hi,
>>>
>>> I'm running version 2.7 on a unix server. I want to create a
>>> custom cache using my own gtf and fasta with gtf2vep.pl
>>> <http://gtf2vep.pl>. This works without problem and also
>>> running VEP seems to go fine. The problem is that, in the
>>> output it seems that the cDNA_position, CDS_position and
>>> Protein_position are correct given my input gtf file but the
>>> calls for Amino_acids and Codons seem completely random. If
>>> I run against the cache retrieved from ensembl these are all
>>> correct. The version of the genome didn't have an effect on
>>> the output, the gtf's haven't changed. The gtf and the fasta
>>> that I'm using for the custom originate from the ensembl
>>> reference so I don't see any reason why the custom cache
>>> shouldn't perform the same way as the reference from ensembl
>>> cache. Could there be bug that somehow messes up the link
>>> between the custom gtf and fasta in my run? Below are the
>>> commands I ran and a snippet of the output's I got.
>>>
>>> Thanks,
>>> Heidi Viitaniemi
>>>
>>> For custom cache I'm running (wrong output for Amino_acids
>>> and Codons)
>>> perl gtf2vep.pl <http://gtf2vep.pl> -i
>>> GasAcu1.67_group_xixflip.gtf -f
>>> gasAcu_group_withoutbac_inv7.fa -d 67 -s
>>> Gasterosteus_aculeatus_XIXflipped_18032013
>>> perl variant_effect_predictor.pl
>>> <http://variant_effect_predictor.pl> -offline 1 -dir
>>> $HOME/.vep -i ens_realigned_AK_F.var.vcf -format vcf -fork 4
>>> -db_version 67 -species
>>> Gasterosteus_aculeatus_XIXflipped_18032013 -numbers
>>> -per_gene -buffer_size 10000 -o
>>> VEP_18032013_exon_pergene_AK_F.var.vcf.txt
>>>
>>> groupXIX_2822477_C/T groupXIX:2822477 T
>>> ENSGACG00000003129 ENSGACT00000004109 Transcript
>>> missense_variant 67 49 17 G/R Gga/Aga - EXON=1/2
>>> groupXIX_2822500_T/C groupXIX:2822500 C
>>> ENSGACG00000003129 ENSGACT00000004109 Transcript
>>> missense_variant 44 26 9 Y/C tAt/tGt - EXON=1/2
>>> groupXIX_2822523_C/T groupXIX:2822523 T
>>> ENSGACG00000003129 ENSGACT00000004109 Transcript
>>> synonymous_variant 21 3 1 R cgG/cgA - EXON=1/2
>>> groupXIX_2822541_T/A groupXIX:2822541 A
>>> ENSGACG00000003129 ENSGACT00000004109 Transcript
>>> 5_prime_UTR_variant 3 - - - - - EXON=1/2
>>>
>>>
>>>
>>> For ensembl cache I'm running (correct output for
>>> Amino_acids and Codons)
>>> perl variant_effect_predictor.pl
>>> <http://variant_effect_predictor.pl> -offline -dir
>>> $HOME/.vep -i ens_realigned_AK_F.var.vcf -format vcf -fork 4
>>> -db_version 69 -species gasterosteus_aculeatus -numbers
>>> -per_gene -buffer_size 10000 -o
>>> ensVEP_18032013_exon_pergene_AK_F.var.vcf.txt
>>>
>>> groupXIX_2822477_C/T groupXIX:2822477 T
>>> ENSGACG00000003129 ENSGACT00000004109 Transcript
>>> missense_variant 67 49 17 A/T Gcg/Acg - EXON=1/2
>>> groupXIX_2822500_T/C groupXIX:2822500 C
>>> ENSGACG00000003129 ENSGACT00000004109 Transcript
>>> missense_variant 44 26 9 D/G gAc/gGc - EXON=1/2
>>> groupXIX_2822523_C/T groupXIX:2822523 T
>>> ENSGACG00000003129 ENSGACT00000004109 Transcript
>>> initiator_codon_variant 21 3 1 M/I atG/atA - EXON=1/2
>>> groupXIX_2822541_T/A groupXIX:2822541 A
>>> ENSGACG00000003129 ENSGACT00000004109 Transcript
>>> 5_prime_UTR_variant 3 - - - - - EXON=1/2
>>>
>>>
>>>
>>> --
>>> ______________________________________________
>>>
>>> Heidi Viitaniemi
>>> PhD student
>>> Division of Genetics and Physiology
>>> Department of Biology
>>> Itäinen Pitkäkatu 4A, 7th floor (Pharmacity)
>>> University of Turku
>>> 20520 Turku
>>>
>>> FINLAND
>>>
>>>
>>> _______________________________________________
>>> Dev mailing list Dev at ensembl.org <mailto:Dev at ensembl.org>
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>>>
>>>
>>>
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>>
>> --
>> ______________________________________________
>>
>> Heidi Viitaniemi
>> PhD student
>> Division of Genetics and Physiology
>> Department of Biology
>> Itäinen Pitkäkatu 4A, 7th floor (Pharmacity)
>> University of Turku
>> 20520 Turku
>>
>> FINLAND
>>
>>
>> _______________________________________________
>> Dev mailing list Dev at ensembl.org <mailto:Dev at ensembl.org>
>> Posting guidelines and subscribe/unsubscribe info:
>> http://lists.ensembl.org/mailman/listinfo/dev
>> Ensembl Blog: http://www.ensembl.info/
>>
>>
>>
>>
>> _______________________________________________
>> Dev mailing listDev at ensembl.org
>> Posting guidelines and subscribe/unsubscribe info:http://lists.ensembl.org/mailman/listinfo/dev
>> Ensembl Blog:http://www.ensembl.info/
>
> --
> ______________________________________________
>
> Heidi Viitaniemi
> PhD student
> Division of Genetics and Physiology
> Department of Biology
> Itäinen Pitkäkatu 4A, 7th floor (Pharmacity)
> University of Turku
> 20520 Turku
>
> FINLAND
>
>
> _______________________________________________
> Dev mailing list Dev at ensembl.org
> Posting guidelines and subscribe/unsubscribe info: http://lists.ensembl.org/mailman/listinfo/dev
> Ensembl Blog: http://www.ensembl.info/
--
______________________________________________
Heidi Viitaniemi
PhD student
Division of Genetics and Physiology
Department of Biology
Itäinen Pitkäkatu 4A, 7th floor (Pharmacity)
University of Turku
20520 Turku
FINLAND
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