Question: Strand Specificity
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gravatar for Aleks Schein
7.2 years ago by
Aleks Schein60
Aleks Schein60 wrote:
Dear All, I am trying to analyze my RNAseq data by TopHat-Cufflinks package based on Galaxy. I used Epicentre ScriptSeq strand-specific library construction protocol, which I assume, produces a second-strand library. When I set FR option to "second strand" and run TopHat and Cufflinks it results in confusing transcript orientation. Cufflinks-assembled transcript with multiple exons are oriented as expected. However, transcripts that entirely reside in one exon/intron or intergenic region are labeled in an opposite way. If TopHat accepted hits track contains negative-strand reads (colored red), transcript is labeled as "positive" and vice versa. An example is shown in the attached screenshot. Both transcripts TCONS_00000014 and TCONS_00000015 were mapped to "+" strand, though reads that represent them had been mapped to "-" strand. Is there a way to solve this issue? Regards, Aleks Schein
rna-seq cufflinks • 1.3k views
ADD COMMENTlink modified 7.2 years ago by Jeremy Goecks2.2k • written 7.2 years ago by Aleks Schein60
0
gravatar for Jeremy Goecks
7.2 years ago by
Jeremy Goecks2.2k
Jeremy Goecks2.2k wrote:
Aleks, This an assumption probably worth verifying before going further. The Tophat documentation may be helpful as well: http://tophat.cbcb.umd.edu/manual.html I haven't heard of or encountered this problem before. Your best bet is probably to email the Tophat/Cufflinks authors and see if they can shed light on your issues: tophat.cufflinks@gmail.com Good luck, J.
ADD COMMENTlink written 7.2 years ago by Jeremy Goecks2.2k
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