<div dir="ltr"><p style="margin:0px;font-size:22px;font-family:Menlo">Dear Jason Stajich,</p>
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<p style="margin:0px;font-size:22px;font-family:Menlo">I am new in BioPerl. I run e-PCR version 2.3.11 on a genomic sequence, and got a output file "seq1.epcr" by following command:</p>
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<p style="margin:0px;font-size:22px;font-family:Menlo">./e-PCR -w9 -f 1 -m5000 test.sts WS240.genomic.fa N=1 G=1 T=3 >seq1.epcr</p>
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<p style="margin:0px;font-size:22px;font-family:Menlo">"seq1.epcr" output looks like:</p>
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<p style="margin:0px;font-size:22px;font-family:Menlo">I FOR_F32H2.2 + 8966315 8966961 647/5000-5000 1 0</p>
<p style="margin:0px;font-size:22px;font-family:Menlo">I FOR_Y54E10BR.d - 3028477 3031091 2615/5000-5000 1 0</p>
<p style="margin:0px;font-size:22px;font-family:Menlo">III FOR_B0280.1.v5 + 7133931 7135112 1182/5000-5000 2 0</p>
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<p style="margin:0px;font-size:22px;font-family:Menlo">Now, I want to extract the amplicon sequences in fasta format from "WS240.genomic.fa" according to STS hits result "seq1.epcr".</p>
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<p style="margin:0px;font-size:22px;font-family:Menlo">However, I want amplicon sequences contains:</p>
<p style="margin:0px;font-size:22px;font-family:Menlo">(1) Only exon sequence</p>
<p style="margin:0px;font-size:22px;font-family:Menlo">(2) If primer hits on non-exon (intron) region, take only exon sequence and write that "forward or reverse" primer hit the intron region.</p>
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<p style="margin:0px;font-size:22px;font-family:Menlo">Could you please tell me how can I use GFF3 annotation file in Bio::Tools::EPCR to extract my amplicon sequences?</p>
<p style="margin:0px;font-size:22px;font-family:Menlo">I have gone through the<span class="" style="white-space:pre"> </span>documentation of "Bio::Tools::EPCR", but I am facing difficult to use it.</p>
<p style="margin:0px;font-size:22px;font-family:Menlo">I would<span class="" style="white-space:pre"> </span>appreciate if you could<span class="" style="white-space:pre"> </span>kindly help me to write a code for it.</p>
<p style="margin:0px;font-size:22px;font-family:Menlo"><br></p><p style="margin:0px;font-size:22px;font-family:Menlo">Note: I have also loaded the "GFF3 and Genomic sequence" in mysql database using "<a href="http://bp_seqfeature_load.pl">bp_seqfeature_load.pl</a>"</p>
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<p style="margin:0px;font-size:22px;font-family:Menlo">Thanks</p>
<p style="margin:0px;font-size:22px;font-family:Menlo">Firoz </p><p style="margin:0px;font-size:22px;font-family:Menlo">New York University, USA</p><div><br></div></div>