Dear all, I am using limma for the analysis of a factorial experiment for the first time. I would greatly appreciate your help re to few questions that I could not understand. The experiment is 2^3 factorial, all the 3 factors at 2 levels each. I have 24 samples. The 3 factors ar below, it is a balanced design genotype<-rep(c(1,2),each=12) ##here 2 represents RAG samples and 1 are WT notch<-rep(rep(1:2,each=2),6)##Here 2 represents DL4 and 1 is OP9 trt<-rep(rep(1:2,2),6)##here 2 represents IL7 treated, while 1 is untreated I define my design matrix as design <- model.matrix(~factor(trt)*factor(notch)*factor(genotype)) The experimenters are interested in genes that are DE for the main effects and the co-regulation of those effects. So I do not define any contrast matrix, is that correct? What I mean to ask is that coefficients estimated by design matrix would suufice the objectives?  Thanks a lot for your help Sincerely, Shaheena [[alternative HTML version deleted]]