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Old 03-21-2016, 07:10 PM   #1
Sentinel156
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Location: Melbourne, Australia

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Default DGE using stranded and unstranded Illumina libraries

Hi all,

I am working with S. pombe doing 100bp single end Illumina HiSeq 2500 RNA-seq. Despite being told the trueseq stranded protocol was used one lot of my libraries appear unstranded (roughly 50/50 split between fwd and rev mapping reads in IGV). The second lot appears stranded as expected (>95% mapping bias to antisense strand in IGV). I'm trying to understand the best way to do read summarisation for these libraries using featurecounts for DGE analysis. I am fearful that the unstranded nature of my first lot of libraries will bias those datasets for genes which have known antisense transcripts. So what is the recommended course of action here? I'm not overly interested in antisense stuff so should i treat everything as unstranded for the featurecounts run?

Thank you in advance.
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Old 03-22-2016, 08:43 AM   #2
fanli
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cross-posted:
https://www.biostars.org/p/182758/
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