Hello,
Sorry for the simple question, but I could not find a straightforward answer to it.
I have RNA-seq data for three different cell lines. Each cell line is treated with three different conditions (Treatment A, Treatment B, and Control). The question I'm trying to address is "How is Treatment B different than Treatment A across the different cell lines?", and I'm using DESeq2 for the analysis.
In order to respond to this question, I believe I have to find the genes whose dysregulations are different between treatment A compared to control and treatment B compared to control. This way, genes that are dysregulated in the same direction with similar effect sizes should not be called differentially expressed and genes that are dysregulated in opposite directions or in the same direction but with differing effect sizes should be called differentially expressed. The primary problem is that I'm not sure about the appropriate design formulae and the contrast I should use for this.
I believe the design ~ Line + Treatment
should be appropriate in this case, but how should I compare the condition effects of the Treatment A compared to control and Treatment B compared to control? Additionally, is it possible to use apeglm shrinkage for this comparison? or am I limited to ashr?
Any help is much Appreciated. Thanks.
Thank you very much for answering my questions! Just one question about your second paragraph:
Isn't this an example of the requirement for blocking? And shouldn't the design
~ Line + Treatment
account for the within-cell line differences? Based on your explanation of how comparing the effects of the treatment A vs control and treatment B vs control is the same as directly comparing treatment A vs treatment B, I believe using a design~ Line + Treatment
followed by extracting the effect for the "Treatment_A_vs_B" should give me what I'm looking for. Am I missing something here?Thanks again for your help!
Oh, wait. I might have misinterpreted your original question. I thought you wanted to know cell-line specific differences, not consistent differences across all cell lines. In that case you can block on line and then do the treatment contrast.
Great! Thank you very much for all your help. Much appreciated!