Hello, I’m working with an RNAseq dataset that looks at plants that are either infected with a fungus or have been left uninfected. I have both male and female genotypes, and those have been cloned, with one clone of each genotype getting inoculated while the other clone serves as the control. I’m also looking at floral and meristematic tissues, but I’m having a hard time figuring out how to use DESeq2 to make all the appropriate comparisons. I’ve seen posts like DESeq2: Paired Test, as well as looking at the vignette, but I don’t understand a few things. Assuming I have a metadata table with sex, condition, and tissue as the columns, with either male or female, infected or uninfected, and flower or meristem in them respectively. If I want to compare male infected flowers to female infected flowers, would the design look like this?
design(~sex + condition + tissue)
Or would it look more like
Or something else entirely. I can’t seem to get my head around how I go about saying I want all the male infected flowers compared to all the female infected flowers, don’t consider other columns like male or female uninfected flowers or any meristem tissues. Additionally, is there a way to specify between doing a paired vs independent comparison? I’d like comparisons like male infected flowers vs male uninfected flowers to be paired since it’s comparing the same genotypes, but comparisons like male infected flower vs female infected flower to be independent since they’re not the same genotypes, and I don’t understand how to distinguish between the two approaches. Thanks very much for any help!