Re: HTSeq-Count

Hi Steve,

thank you very much for your help.

Which tool for DE did you use?
I used edger, however I´ve read that edgeR and DESeq2 might be overstringent for RIP-Seq (f.e. RIP-Seeker package Paper, Supplement).

Best wishes,
Julia

-----Ursprüngliche Nachricht-----
Von: [email protected] [mailto:[email protected]] Im Auftrag von Steve Lianoglou
Gesendet: Freitag, 5. September 2014 18:35
An: Julia [guest]
Cc: [email protected] list; Pickl, Julia
Betreff: Re: [BioC] HTSeq-Count

Hi Julia,

On Fri, Sep 5, 2014 at 7:41 AM, Julia [guest] <[email protected]> wrote:
> Hi all,
> I am new to the field of seq and performed a RIP-Seq experiment using HTSeq count as counter.
> I get now the following (using union, but doesn´t look better for interesection_strict):
> __no_feature    1503377
> __ambiguous     490772
> __too_low_aQual 0
> __not_aligned   0
> __alignment_not_unique  5277314
>
> When I sum up counts for all genes, I get 3227845.
>
> The number for __no_feature, __ambiguous, __alignment_not_unique look very high.
>
> Does somebody have an idea for that?

While I haven't worked with RIP-seq data myself, I do have some experience with HITS-CLIP and PAR-CLIP, which I believe are quite similar (at least in principle) -- these experiments must incredibly difficult to pull off, however, because I'd say most of these types of datasets that came my way were notoriously/incredibly noisy.

This is just to say the problem you are seeing may not be an informatics problem, and could (quite possibly) be an experimental one.

-steve

--
Steve Lianoglou
Computational Biologist
Genentech

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