Hello, I am using DESeq2 on a project, where I tested various extraction volumes and types and got amplicon data out. So for instance, I have two extraction volumes 100 Liters and 20 Liters and then I have two extraction methods PowerWater and PowerSoil. I want to compare the resulting microbiomes. So at first I setup my experiment as: equation ~ExtractionType + Volume; then changed it to equation ~Volume + ExtractionType (I am getting basically the same PCA results both ways). I am wondering if I should change it to equation ~ExtractionType + Volume + ExtractionType:Volume so that I have all possible varieties in the equation?

I also have 2 different sites being tested, but I was going to set up different experiments for them and just pull out one site set up deseq2 and then do the other experiment.

A few other issues I am having, my Volume is being treated as numeric, even when I turned it into a factor, should I change it to be words in order to force it to be a character instead of numeric or keep it as numeric?

Also I need to test the hypothesis that the micrbiome is the same at both Volumes or Extraction methods. Can you please tell me how I would do so in DESeq?

Thank you so much for your time!

The choice of design and which hypotheses you test is up to you. Unfortunately I can’t provide this level of support on the support site, I have to limit to software usage questions.

Regarding not being able to convert numeric to factor can you post your code? How do you determine it is not being received as factor?