I have RNA-seq data from nuclear (Nuc) and cytoplasmic (cyto) fractions of both undifferentiated (UD) and differentiated (DF) cells (I have triplicates of all conditions).
Essentially I have UDNuc1, UDNuc2, UDNuc3, UDcyto1, UDcyto2, UDcyto3, DFNuc1, DFNuc2, DFNuc3, DFcyto1, DFcyto2, DFcyto3.
I have run DESeq2 on UD nuc vs UD cyto samples and DF nuc vs DF cyto samples separately but I am trying to compare the RATIOS of UD vs DF. For example I want DESeq2 to tell me if there is a difference between (UDnuc/UDcyto) and (DFnuc/DFcyto).
I was trying to just manually (in R) calculate the ratios first then run that through DESeq2 but I've been having problems with non-integer numbers and NA's. Currently this is the design I have and I'm sure I just need to change the condition but I want to make sure I do it correctly.
condition <- factor(c(rep("UD", 3), rep("DF", 3))) coln <- colnames(countdata) coldata <- data.frame(row.names=coln, condition) dds <- DESeqDataSetFromMatrix(countData=countdata, colData=coldata, design=~condition)
Any advice on how to set up my design would be greatly appreciated!
