hi Yoong,

On Wed, Nov 20, 2013 at 5:56 PM, FeiYian Yoong [guest] <
guest@bioconductor.org> wrote:

>
> Hi there,
>
> I am wondering:
>
> i) When do you perform count outlier detection step? Do you do it after
> filtering/differential expression analysis OR before both steps?
>

Count outlier detection is performed before the p-value adjustment, which
is the point where it would make a difference before or after. So it does
not count in multiple testing to the number of tests performed.
​​

>
> ii) It doesn't seem like DESeq2 take into account normalizing samples
> against endogenous or consistently expressed genes for differential
> expression analysis. Do you recommend that?
>

​DESeq2 uses the median ratio method described in Anders & Huber 2010 over
all genes. It does not take into account any prior information about the
genes. However, if you know a certain subset of genes are not
differentially expressed, you could use the following code to produce size
factors over these genes, indexed by 'nonDEgenes':

sf <- estimateSizeFactorsForMatrix​( counts(dds)[ nonDEgenes , ] )
​sizeFactors(dds) <- sf​

​hope this helps,

Mike​





>
>
> Thank you in advance!!
>
> Yoong
>
>  -- output of sessionInfo():
>
> NA
>
> --
> Sent via the guest posting facility at bioconductor.org.
>

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