Hi, I would be grateful for some input on using edgeR for small RNA sequence data. I have been testing edgeR on a set of miRNA data (3 groups with n=10, 15 and 15). After removing genes that are not expressed at >= 0.2 cpm in >= 5 samples I have ~600 rows left. I tried calculating the tagwise dispersion estimate with: cds1 <- estimateTagwiseDisp(cds1, prior.n=2, trend=TRUE, prop.used=0.1, grid=FALSE) Increasing the prior to e.g. 10 gives more differentially expressed genes that do not look bad. Decreasing the prior to 0 leaves me with extremely few differentially expressed genes that are mainly variance outliers. I guess that miRNA data is likely to behave differently from mRNA data since there are so few genes (but still a very large dynamic range). Is it possible that I am over-fitting the estimate? Would you recommend changing any other parameters? Best regards, Helena _________________________________ Helena Persson, PhD Karolinska Institutet Dept of Biosciences and Nutrition Hälsovägen 7-9 SE-141 83 Huddinge Sweden Helena.Persson@ki.se tel. +46-(0)8-52481058 [[alternative HTML version deleted]]