Question: How To Normalization Multiple Groups At Once: Edger Or Deseq
0
gravatar for Rm
6.5 years ago by
Rm7.8k
Danville, PA
Rm7.8k wrote:

I have three groups (A, B, C) to compare each with many samples: My goal is to normalize all groups as a whole and then get differential expression (DE) list for each pair.

My attempts so far with edgeR:

if i define my groups [ group <- c(rep("A",68),rep("B",65),rep("C",70)) ] before calcNormFactors and common estimateCommonDisp and exporting normalized values. It looks like normalization is occurring within each group (which i am not looking for)

what I tried is define entire samples as one group [ group <- c(rep("A",203)) ] . I could get the normalized values: but my difficulty now it is not able to perform exactTest as group has only one entity.

After normalization I redefined the groups as in my first case still unable to perform the exactTest.

I really appreciate if you can suggest to over come this.... I am open to using other tools if they support this.

multiple edger • 3.0k views
ADD COMMENTlink modified 6.5 years ago by Mikael Huss4.6k • written 6.5 years ago by Rm7.8k

Can you clarify a bit what you mean by this sentence: "After normalization I redefined the groups as in my first case still unable to perform the exactTest."? Did you normalize considering everything as one group, and then do something like d$samples$group <- c(rep("A",68),rep("B",65),rep("C",70)) (assuming your DGEList object is called d)? And in what way were you unable to perform the exactTest? Was there an error message?

ADD REPLYlink written 6.5 years ago by Mikael Huss4.6k

As Mikael's prompt for further information is alluding to, it would be much more helpful if you pasted in the code you used for this analysis -- it won't be much. You can then inline in comments (or break code into sections) and ask specific questions about parts of it. This would help us help you a lot better.

Also, what makes you say that "it looks like normalization is occurring within each group"

ADD REPLYlink written 6.5 years ago by Steve Lianoglou5.0k

Thanks @Mikael; it worked after following your suggestion "d$samples$group" . Can you post it as answer i will accept it..

ADD REPLYlink modified 6.5 years ago • written 6.5 years ago by Rm7.8k
1
gravatar for Mikael Huss
6.5 years ago by
Mikael Huss4.6k
Stockholm
Mikael Huss4.6k wrote:

Did you normalize considering everything as one group, and then do something like d$samples$group <- c(rep("A",68),rep("B",65),rep("C",70))?

I suppose it would also be possible to normalize everything as one group (again) and then set up a design matrix and contrasts that would describe the comparison you are looking for.

ADD COMMENTlink written 6.5 years ago by Mikael Huss4.6k
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