[Bioc-sig-seq] RNASeq, differential expression between group, and large variance within groups

[Steve Lianoglou]

Hi,

Just to add to what Mads said -- most Tag sequencing protocols involve
a pcr amplification step, which might be causing the result you see
here. Unfortunately, unlike "normal" rna-seq, with tag sequencing, you
can't use information like coverage in the local neighborhood of your
"rogue" tag to help flag pcr amplification artifacts.

> Depending on the cutting enzyme, is it the most 3' canonical, or could it be
> caused by incomplete digestion?

I would guess that incomplete digestion would happen across the board, though.

Assuming you are just interested in using the tag-seq data to identify
overall gene expression, wouldn't it make sense to sum all of the tag
counts over your gene (whether they are 3' canonical or otherwise),
and feed those counts into edgeR/DESeq, etc. for expression analysis
anyway?

-- 
Steve Lianoglou
Graduate Student: Computational Systems Biology
 | Memorial Sloan-Kettering Cancer Center
 | Weill Medical College of Cornell University
Contact Info: http://cbio.mskcc.org/~lianos/contact