<div dir="ltr">Thank you very much for your response. Yes, that is the issue, and after it had been fixed, the file went through the program just fine.<div><br></div><div>Thanks,</div><div><br></div><div>Xin</div></div><div class="gmail_extra"><br><div class="gmail_quote">On Tue, Aug 25, 2015 at 12:51 PM, Carson Holt <span dir="ltr"><<a href="mailto:carsonhh@gmail.com" target="_blank">carsonhh@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 style="word-wrap:break-word">You have edited the file in a way that broke it. Gene maker-scaffold10370-exonerate_est2genome-gene-0.0 previously had two separate transcript sequences (mRNA-1 and mRNA-2). You have deleted mRNA-1, but some of it’s children exons still exist. Notice that some exons have two parents, and you have deleted one of the parents without removing the parent relationship.<div><br></div><div>—Carson</div><div><br><div><br></div><div><br><div><blockquote type="cite"><div><div class="h5"><div>On Aug 20, 2015, at 9:37 AM, Xin Huang <<a href="mailto:xh33@georgetown.edu" target="_blank">xh33@georgetown.edu</a>> wrote:</div><br></div></div><div><div><div class="h5"><div dir="ltr">Dear Developer,<div><br></div><div>I was trying to use a mixed GFF3 file to do some analysis, which includes genome-level gene predictions and manually curated annotations. I replaced the gene prediction by Maker annotations where appropriate. And a sample of the blended GFF3 file is as follows:</div><div><br></div><div><div>scaffold10370<span style="white-space:pre-wrap"> </span>GLEAN<span style="white-space:pre-wrap"> </span>mRNA<span style="white-space:pre-wrap"> </span>3918<span style="white-space:pre-wrap"> </span>18376<span style="white-space:pre-wrap"> </span>0.920154<span style="white-space:pre-wrap"> </span>+<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=CCG000529.1;source_id=Aalb_GLEAN_10002648;</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>GLEAN<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>3918<span style="white-space:pre-wrap"> </span>3983<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>+<span style="white-space:pre-wrap"> </span>0<span style="white-space:pre-wrap"> </span>Parent=CCG000529.1;</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>GLEAN<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>4521<span style="white-space:pre-wrap"> </span>4598<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>+<span style="white-space:pre-wrap"> </span>0<span style="white-space:pre-wrap"> </span>Parent=CCG000529.1;</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>GLEAN<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>17407<span style="white-space:pre-wrap"> </span>17516<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>+<span style="white-space:pre-wrap"> </span>0<span style="white-space:pre-wrap"> </span>Parent=CCG000529.1;</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>GLEAN<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>17588<span style="white-space:pre-wrap"> </span>17767<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>+<span style="white-space:pre-wrap"> </span>1<span style="white-space:pre-wrap"> </span>Parent=CCG000529.1;</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>GLEAN<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>18238<span style="white-space:pre-wrap"> </span>18376<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>+<span style="white-space:pre-wrap"> </span>1<span style="white-space:pre-wrap"> </span>Parent=CCG000529.1;</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>gene<span style="white-space:pre-wrap"> </span>32452<span style="white-space:pre-wrap"> </span>54508<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0;Name=AAEL004146</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>mRNA<span style="white-space:pre-wrap"> </span>32452<span style="white-space:pre-wrap"> </span>54508<span style="white-space:pre-wrap"> </span>44204<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0;Name=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2;aED=0.00;eAED=0.00;qI=376|1|1|1|0|0|5|316|548</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>exon<span style="white-space:pre-wrap"> </span>32452<span style="white-space:pre-wrap"> </span>33011<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2:exon:15;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>exon<span style="white-space:pre-wrap"> </span>35140<span style="white-space:pre-wrap"> </span>35388<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1:exon:12;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1,maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>exon<span style="white-space:pre-wrap"> </span>35455<span style="white-space:pre-wrap"> </span>36119<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1:exon:11;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1,maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>exon<span style="white-space:pre-wrap"> </span>36443<span style="white-space:pre-wrap"> </span>36777<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1:exon:10;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1,maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>exon<span style="white-space:pre-wrap"> </span>53979<span style="white-space:pre-wrap"> </span>54508<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1:exon:9;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1,maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>five_prime_UTR<span style="white-space:pre-wrap"> </span>54133<span style="white-space:pre-wrap"> </span>54508<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2:five_prime_utr;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>53979<span style="white-space:pre-wrap"> </span>54132<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>0<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2:cds;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>36443<span style="white-space:pre-wrap"> </span>36777<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>2<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2:cds;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>35455<span style="white-space:pre-wrap"> </span>36119<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>0<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2:cds;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>35140<span style="white-space:pre-wrap"> </span>35388<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>1<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2:cds;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>CDS<span style="white-space:pre-wrap"> </span>32768<span style="white-space:pre-wrap"> </span>33011<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>1<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2:cds;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div><div>scaffold10370<span style="white-space:pre-wrap"> </span>maker<span style="white-space:pre-wrap"> </span>three_prime_UTR<span style="white-space:pre-wrap"> </span>32452<span style="white-space:pre-wrap"> </span>32767<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>-<span style="white-space:pre-wrap"> </span>.<span style="white-space:pre-wrap"> </span>ID=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2:three_prime_utr;Parent=maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-2</div></div><div><br></div>This file does not pass the GFF3 online validator from GenomeTools with the following error message:<div><br>GenomeTools error: Parent "maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1" on line 1456 in file "/var/www/servers/<a href="http://genometools.org/htdocs/cgi-bin/gff3/Aalbo.gff" target="_blank">genometools.org/htdocs/cgi-bin/gff3/Aalbo.gff</a>" was not defined (via "ID=")</div><div><br></div><div>And when I tried to use gff3toGenePred to convert GFF3 to the refflat format, I got the following parsing errors (samples):</div><div><br></div>Can't find annotation record "maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1" referenced by "maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1:exon:12" Parent attribute<br><br>Can't find annotation record "maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1" referenced by "maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1:exon:11" Parent attribute<br><br>Can't find annotation record "maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1" referenced by "maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1:exon:10" Parent attribute<br><br>Can't find annotation record "maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1" referenced by "maker-scaffold10370-exonerate_est2genome-gene-0.0-mRNA-1:exon:9" Parent attribute<br><br>Can't find annotation record "maker-scaffold1193-exonerate_est2genome-gene-2.0-mRNA-2" referenced by "maker-scaffold1193-exonerate_est2genome-gene-2.0-mRNA-1:exon:114" Parent attribute<br><br>Can't find annotation record "maker-scaffold16459-exonerate_est2genome-gene-0.0-mRNA-1" referenced by "maker-scaffold16459-exonerate_est2genome-gene-0.0-mRNA-1:exon:118" Parent attribute<br><br>Can't find annotation record "maker-scaffold16459-exonerate_est2genome-gene-0.0-mRNA-1" referenced by "maker-scaffold16459-exonerate_est2genome-gene-0.0-mRNA-1:exon:119" Parent attribute<br><br>Can't find annotation record "maker-scaffold16459-exonerate_est2genome-gene-0.0-mRNA-1" referenced by "maker-scaffold16459-exonerate_est2genome-gene-0.0-mRNA-1:exon:120" Parent attribute<br><br>Can't find annotation record "maker-scaffold16459-exonerate_est2genome-gene-0.0-mRNA-1" referenced by "maker-scaffold16459-exonerate_est2genome-gene-0.0-mRNA-1:exon:121" Parent attribute<div><br></div><div>What could have been the issue in the annotation file or the way I used the file? Any feedback will be highly appreciated!</div><div><br></div><div>Sincerely,</div><div><br></div><div>Xin Huang</div></div></div></div>
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