<div dir="ltr">Thank you, Fiona. That is very helpful!<div><br></div><div>Dan</div></div><div class="gmail_extra"><br><div class="gmail_quote">On Tue, Aug 4, 2015 at 10:37 AM, Fiona Cunningham <span dir="ltr"><<a href="mailto:fiona@ebi.ac.uk" target="_blank">fiona@ebi.ac.uk</a>></span> wrote:<br><blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex"><div dir="ltr">Hi Dan,<div><br></div><div>Thanks for getting in touch. The VEP considers each variant separately, even if they are in the same codon. This is because the variants may be on different strands. You can add information on this using a plugin e.g. <a href="https://github.com/ensembl-variation/VEP_plugins/blob/master/SameCodon.pm" target="_blank">https://github.com/ensembl-variation/VEP_plugins/blob/master/SameCodon.pm</a></div><div><br></div><div>See more info here: <a href="http://www.ensembl.org/info/docs/tools/vep/script/vep_plugins.html" target="_blank">http://www.ensembl.org/info/docs/tools/vep/script/vep_plugins.html</a></div></div><div class="gmail_extra"><br clear="all"><div><div>Fiona<br>-----------------------------------------------------------------<br>Fiona Cunningham, PhD<br>Variation Annotation Coordinator, <br>European Bioinformatics Institute (EMBL-EBI)<br><a href="http://www.ensembl.org" target="_blank">www.ensembl.org</a> || <a href="http://www.lrg-sequence.org" target="_blank">www.lrg-sequence.org</a></div></div>
<br><div class="gmail_quote"><span class="">On 30 July 2015 at 23:00, Dan Sun <span dir="ltr"><<a href="mailto:meredithfy@gmail.com" target="_blank">meredithfy@gmail.com</a>></span> wrote:<br></span><blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex"><div dir="ltr"><span class="">Hi Will,<div><br></div><div>Thanks again! I have another minor bug to report.</div><div><br></div><div>For mutations in the same codon, VEP annotates them separately. This could sometimes cause problems. The following is the VEP output for two mutations in the same codon:</div><div><br></div><div>NW_005081561.1_649917_G/A<span style="white-space:pre-wrap"> </span>NW_005081561.1:649917<span style="white-space:pre-wrap"> </span>A<span style="white-space:pre-wrap"> </span>102066196<span style="white-space:pre-wrap"> </span>XM_005485125.1<span style="white-space:pre-wrap"> </span>Transcript<span style="white-space:pre-wrap"> </span>missense_variant<span style="white-space:pre-wrap"> </span>6577<span style="white-space:pre-wrap"> </span>6577<span style="white-space:pre-wrap"> </span>2193<span style="white-space:pre-wrap"> </span>D/N<span style="white-space:pre-wrap"> </span>Gat/Aat<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>IMPACT=MODERATE;STRAND=1</div><div>NW_005081561.1_649918_A/G<span style="white-space:pre-wrap"> </span>NW_005081561.1:649918<span style="white-space:pre-wrap"> </span>G<span style="white-space:pre-wrap"> </span>102066196<span style="white-space:pre-wrap"> </span>XM_005485125.1<span style="white-space:pre-wrap"> </span>Transcript<span style="white-space:pre-wrap"> </span>missense_variant<span style="white-space:pre-wrap"> </span>6578<span style="white-space:pre-wrap"> </span>6578<span style="white-space:pre-wrap"> </span>2193<span style="white-space:pre-wrap"> </span>D/G<span style="white-space:pre-wrap"> </span>gAt/gGt<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>IMPACT=MODERATE;STRAND=1</div><div><br></div><div>However, instead of GAT -> AAT or GAT -> GGT, the true mutation is GAT -> AGT. The amino acid changes from D to S, not to N or G. I think an output like this might make more sense:</div><div><br></div><div>NW_005081561.1_649917_GA/AG<span style="white-space:pre-wrap"> </span>NW_005081561.1:649917-649918<span style="white-space:pre-wrap"> </span>AG<span style="white-space:pre-wrap"> </span>102066196<span style="white-space:pre-wrap"> </span>XM_005485125.1<span style="white-space:pre-wrap"> </span>Transcript<span style="white-space:pre-wrap"> </span>missense_variant<span style="white-space:pre-wrap"> </span>6577-6578<span style="white-space:pre-wrap"> </span>6577-6578<span style="white-space:pre-wrap"> </span>2193<span style="white-space:pre-wrap"> </span>D/S<span style="white-space:pre-wrap"> </span>Gat/AGt<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>IMPACT=MODERATE;STRAND=1</div><div><br></div><div>Thanks,</div></span><div>Dan</div></div><div><div class="h5"><div><div><div class="gmail_extra"><br><div class="gmail_quote">On Thu, Jul 30, 2015 at 10:21 AM, Dan Sun <span dir="ltr"><<a href="mailto:meredithfy@gmail.com" target="_blank">meredithfy@gmail.com</a>></span> wrote:<br><blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex"><div dir="ltr"><div>Hi Will,</div><div><br></div>Thank you! It works like a charm. <div><br></div><div>Have a great day!</div><span><font color="#888888"><div><br></div><div>Dan</div><div><br></div></font></span></div><div><div><div class="gmail_extra"><br><div class="gmail_quote">On Thu, Jul 30, 2015 at 5:49 AM, Will McLaren <span dir="ltr"><<a href="mailto:wm2@ebi.ac.uk" target="_blank">wm2@ebi.ac.uk</a>></span> wrote:<br><blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex"><div dir="ltr">Hi Dan,<div><br>Thanks for the report, we are still working on ironing out some issues in the GFF parser.</div><div><br></div><div>I've added some fixes to the release/81 version of <a href="http://gtf2vep.pl" target="_blank">gtf2vep.pl</a> which should correct the problems you are seeing.</div><div><br></div><div>Regards</div><span><font color="#888888"><div><br></div><div>Will</div></font></span></div><div><div><div class="gmail_extra"><br><div class="gmail_quote">On 29 July 2015 at 22:21, Dan Sun <span dir="ltr"><<a href="mailto:meredithfy@gmail.com" target="_blank">meredithfy@gmail.com</a>></span> wrote:<br><blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex"><div dir="ltr">Hi Will and Christian,<div><br></div><div>Thank you both for your help.</div><div><br></div>I have an additional question. Once I annotated my vcf file using your cache, I notice non-coding variants are marked "intergenic variant" instead of something like "non coding exon variant". For example, NW_005081553.1: 4008346G->T is a variant located in an exon of non-coding transcripts of gene KHDRBS2 (XR_270793.1, XR_270792.1, XR_270795.1, XR_270797.1, XR_270794.1). You have any ideas about how to improve the annotation of SNPs in exons of non-coding genes for this species? You can find these non-coding transcripts in the GFF3 file you downloaded from NCBI.<div><br></div><div>Thanks! </div><div><br></div><div>Best,</div><div>Dan</div></div><div class="gmail_extra"><div><div><br><div class="gmail_quote">On Tue, Jul 28, 2015 at 5:52 AM, Christian Cole (Staff) <span dir="ltr"><<a href="mailto:C.Cole@dundee.ac.uk" target="_blank">C.Cole@dundee.ac.uk</a>></span> wrote:<br><blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex">
<div style="word-wrap:break-word;color:rgb(0,0,0);font-size:16px;font-family:Cochin,sans-serif">
<div>
<div>
<div>Sorry, I couldn't leave this alone. I don't think I've done enough coding lately ;)</div>
<div><br>
</div>
<div>You can shorten it a fair bit further with the magic -a (auto-split) and -p (auto-print) switches:</div>
<div>gzip -dc 44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa.gz | perl -F'/\|/' -lape 's/^>.*/>$F[3]/' > 44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa</div>
<div>
<div></div>
</div>
</div>
</div>
<div><br>
</div>
<div>-a splits each line by the pattern given by -F (whitespace by default) and puts it into @F</div>
<div>-p puts while{<>} { print } around your code</div>
<div><br>
</div>
<div>Using substitution rather than an if() simplifies the defline fix. Although, it's a lot less legible.</div>
<div><br>
</div>
<div>OK. I feel better now...</div>
<div>Cheers,</div>
<div><br>
</div>
<div>Chris</div>
<div><br>
</div>
<span>
<div style="font-family:Calibri;font-size:12pt;text-align:left;color:black;BORDER-BOTTOM:medium none;BORDER-LEFT:medium none;PADDING-BOTTOM:0in;PADDING-LEFT:0in;PADDING-RIGHT:0in;BORDER-TOP:#b5c4df 1pt solid;BORDER-RIGHT:medium none;PADDING-TOP:3pt"><span>
<span style="font-weight:bold">From: </span><<a href="mailto:dev-bounces@ensembl.org" target="_blank">dev-bounces@ensembl.org</a>> on behalf of Will McLaren<br>
<span style="font-weight:bold">Reply-To: </span>Ensembl developers list<br>
</span><span style="font-weight:bold">Date: </span>Tuesday, 28 July 2015 10:16<div><div><br>
<span style="font-weight:bold">To: </span>Ensembl developers list<br>
<span style="font-weight:bold">Subject: </span>Re: [ensembl-dev] Request to add one species to VEP pre-built cache<br>
</div></div></div><div><div>
<div><br>
</div>
<blockquote style="BORDER-LEFT:#b5c4df 5 solid;PADDING:0 0 0 5;MARGIN:0 0 0 5">
<div>
<div>
<div dir="ltr">Thanks Chris - always good to shorten one-liners.
<div><br>
</div>
<div>And you're correct, the space is not intentional; the command should be:</div>
<br>
gzip -dc 44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa.gz | perl -lne 'if(/^\>/) { $id = (split /\|/, $_)[3]; print ">$id";} else {print}' > 44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa
<div><br>
</div>
<div>Regards<br>
<br>
Will</div>
</div>
<div class="gmail_extra"><br>
<div class="gmail_quote">On 28 July 2015 at 10:09, Christian Cole (Staff) <span dir="ltr">
<<a href="mailto:C.Cole@dundee.ac.uk" target="_blank">C.Cole@dundee.ac.uk</a>></span> wrote:<br>
<blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex">
<div style="word-wrap:break-word;color:rgb(0,0,0);font-size:16px;font-family:Cochin,sans-serif">
<div>
<div>
<div>Hi Will,</div>
<div><br>
</div>
<div>Just a quick tip. Using the perl -n switch avoids 'while(<>) { }' and -l switch avoids having to terminate print statements with '\n'. So your code can be tidied up a touch with:</div>
<div>gzip -dc 44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa.gz | perl -lne 'if(/^\>/) { $id = (split /\|/, $_)[3]; print "> $id";} else {print}' > 44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa</div>
<div><br>
</div>
<div>
<div></div>
</div>
</div>
</div>
<div>Also, is the space in '> $id' intentional? That's not typical behaviour for fasta files.</div>
<div>Cheers,</div>
<div><br>
</div>
<div>Chris</div>
<div><br>
</div>
<span>
<div style="font-family:Calibri;font-size:12pt;text-align:left;color:black;BORDER-BOTTOM:medium none;BORDER-LEFT:medium none;PADDING-BOTTOM:0in;PADDING-LEFT:0in;PADDING-RIGHT:0in;BORDER-TOP:#b5c4df 1pt solid;BORDER-RIGHT:medium none;PADDING-TOP:3pt">
<span style="font-weight:bold">From: </span><<a href="mailto:dev-bounces@ensembl.org" target="_blank">dev-bounces@ensembl.org</a>> on behalf of Will McLaren<br>
<span style="font-weight:bold">Reply-To: </span>Ensembl developers list<br>
<span style="font-weight:bold">Date: </span>Monday, 27 July 2015 17:27<br>
<span style="font-weight:bold">To: </span>Ensembl developers list<br>
<span style="font-weight:bold">Subject: </span>Re: [ensembl-dev] Request to add one species to VEP pre-built cache<br>
</div>
<div>
<div>
<div><br>
</div>
<blockquote style="BORDER-LEFT:#b5c4df 5 solid;PADDING:0 0 0 5;MARGIN:0 0 0 5">
<div>
<div>
<div dir="ltr">Hi Dan,
<div><br>
</div>
<div>We have in fact just updated our GTF converter script to support GFF too (get the new release, 81, for this capability).</div>
<div><br>
</div>
<div>However, giving it a go just now with that file I noticed the FASTA file supplied doesn't play nicely with our indexer, so I tweaked the FASTA to get it to run. Long story short, here's the cache:</div>
<div><br>
</div>
<div><a href="https://dl.dropboxusercontent.com/u/12936195/zonotrichia_albicollis.tar.gz" target="_blank">https://dl.dropboxusercontent.com/u/12936195/zonotrichia_albicollis.tar.gz</a><br>
</div>
<div><br>
</div>
<div>And here's the long story, i.e. what I did to generate it if you want to do the same:</div>
<div><br>
</div>
<div>wget <a href="ftp://ftp.ncbi.nlm.nih.gov/genomes/Zonotrichia_albicollis/GFF/ref_Zonotrichia_albicollis-1.0.1_scaffolds.gff3.gz" target="_blank">
ftp://ftp.ncbi.nlm.nih.gov/genomes/Zonotrichia_albicollis/GFF/ref_Zonotrichia_albicollis-1.0.1_scaffolds.gff3.gz</a><br>
</div>
<div>wget <a href="ftp://ftp.ncbi.nlm.nih.gov/genomes/Zonotrichia_albicollis/CHR_Un/44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa.gz" target="_blank">
ftp://ftp.ncbi.nlm.nih.gov/genomes/Zonotrichia_albicollis/CHR_Un/44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa.gz</a><br>
</div>
<div>gzip -dc 44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa.gz | perl -e 'while(<>) { if(/^\>/) { $id = (split /\|/, $_)[3]; print "> $id\n";} else {print}}' > 44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa</div>
<div>perl <a href="http://gtf2vep.pl" target="_blank">gtf2vep.pl</a> -i ref_Zonotrichia_albicollis-1.0.1_scaffolds.gff3.gz -fasta 44394_ref_Zonotrichia_albicollis-1.0.1_chrUn.fa -species zonotrichia_albicollis</div>
<div><br>
</div>
<div>Then run the VEP as follows:</div>
<div><br>
</div>
<div>perl <a href="http://variant_effect_predictor.pl" target="_blank">variant_effect_predictor.pl</a> -offline -species zonotrichia_albicollis -i variants.vcf</div>
<div><br>
</div>
<div>Regards</div>
<div><br>
</div>
<div>Will McLaren</div>
<div>Ensembl Variation</div>
<div><br>
</div>
<div><br>
</div>
<div><br>
</div>
</div>
<div class="gmail_extra"><br>
<div class="gmail_quote">On 27 July 2015 at 16:49, Dan Sun <span dir="ltr"><<a href="mailto:meredithfy@gmail.com" target="_blank">meredithfy@gmail.com</a>></span> wrote:<br>
<blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex">
<div dir="ltr">Hi,
<div><br>
</div>
<div>I was trying to build a cache from GTF for white-throated sparrow by myself following the tutorial, but was not successful. If possible, could you please add this species to the download list? I would really appreciate that! </div>
<div><br>
</div>
<div>You may download the GFF3 annotation for this species from NCBI ftp (<a href="ftp://ftp.ncbi.nlm.nih.gov/genomes/Zonotrichia_albicollis/GFF/ref_Zonotrichia_albicollis-1.0.1_scaffolds.gff3.gz" target="_blank">ftp://ftp.ncbi.nlm.nih.gov/genomes/Zonotrichia_albicollis/GFF/ref_Zonotrichia_albicollis-1.0.1_scaffolds.gff3.gz</a>)
and convert it to GTF.</div>
<div><br>
</div>
<div>Thank you very much!<span><font color="#888888"><br clear="all">
<div><br>
</div>
-- <br>
<div>
<div dir="ltr">Dan <br>
</div>
</div>
</font></span></div>
</div>
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<br></blockquote></div><br><br clear="all"><div><br></div></div></div><span>-- <br><div><div dir="ltr">Dan Sun <br>Graduate student of Bioinformatics<br>School of Biology<br>Georgia Institute of Technology<br></div></div>
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<br></blockquote></div><br><br clear="all"><div><br></div>-- <br><div class="gmail_signature"><div dir="ltr">Dan Sun <br>Graduate student of Bioinformatics<br>School of Biology<br>Georgia Institute of Technology<br></div></div>
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