Difference in differential gene expression

Question

[Marcel]

Hello,

I have RNA-seq data from 4 different conditions. Basically, it's wild type, knockout, wild type under heatshock and knockout under heatshock. I want to find whether there is a significant change between the wild type and the knockout regarding the up/down regulation of genes under heatshock conditions.

So far, I have analyzed the pairwise differential expressions using DESeq2. What I can now of course do, is to look whether the significantly differentially expressed genes upon heatshock in the wild type are also significantly up/down in the knockout. But it's not exactly what I want. Basically what I what to know is whether there's a significant change in the change upon heatshock between wild type and knockout.

I have so far not found a way to do this. One rather naive idea I had is to run DESeq2 with the difference in readcounts upon heatshock rather than the actual readcounts for each gene. Is this a good idea or there any other or better ways to do this?

[ATpoint]

How did you setup your design parameter? Please show this and the colData. One could do this probably with interactions but it depends on how you encoded your groups.

[Marcel]

I'm using DESeqDataSetFromHTSeqCount. For my pairwise comparisons, I had sampleTables like this:

sample  file    group   replicate
WT_1_NHS    WT1.readcounts.tsv  WT_NHS  1
WT_2_NHS    WT2.readcounts.tsv  WT_NHS  2
WT_3_NHS    WT3.readcounts.tsv  WT_NHS  3
KO_1_NHS    KO1.readcounts.tsv  KO_NHS  1
KO_2_NHS    KO2.readcounts.tsv  KO_NHS  2
KO_3_NHS    KO3.readcounts.tsv  KO_NHS  3

(this one was for WT non-heat shock vs. knockout non-heat shock. My tables for the other comparisons look analogously). The design parameter is "~group". I'll have a closer look at the interactions function of DESeq2 then.

[swbarnes2]

Have you read the vignette? Did you look at the section on interactions?

[Marcel]

Yes, I did look at the vignette. I guess, I misunderstood the interactions section. Thanks for pointing me in that direction.