Dear BioC, I am tring to use Limma package in order to analyze miRNA expression. I would like to identify differentially expressed miRNA; my experimental design encompasses dye swap, and different technical replicates: REF expA1 REF expB1 REF expC1 the corensponing dye-swap expA1 REF expB1 REF expC1 REF REF expA2 REF expB2 REF expC2 the corensponing dye-swap expA2 REF expB2 REF expC2 REF REF expA3 REF expB3 REF expC3 the corensponing dye-swap expA3 REF expB3 REF expC3 REF I how can I set the design matrix for linear model in order to identify differentially regulated genes in respect to the comaparison between A vs B, A vs C and B vs C. I tried with fit <- lmFit(MA, design=c(1,2,3,-1,-2,-3,1,2,3,-1,-2,-3,1,2,3,-1,-2,-3)) but I get results that are too much different for t-test among conditions, so I think it is wrong. thank you Claudio ------------------------------------------------------ Passa a Infostrada. ADSL e Telefono senza limiti e senza canone Telecom http://click.libero.it/infostrada

Quoting "claudio.is at libero.it" <claudio.is at="" libero.it="">: > Dear BioC, > > I am tring to use Limma package in order to analyze miRNA > expression. I would like to identify differentially expressed miRNA; > my experimental design encompasses dye swap, and different > technical replicates: > > REF expA1 > REF expB1 > REF expC1 > the corensponing dye-swap > expA1 REF > expB1 REF > expC1 REF > > REF expA2 > REF expB2 > REF expC2 > the corensponing dye-swap > expA2 REF > expB2 REF > expC2 REF > > REF expA3 > REF expB3 > REF expC3 > the corensponing dye-swap > expA3 REF > expB3 REF > expC3 REF > > I how can I set the design matrix for linear model in order to > identify differentially regulated genes in respect to the > comaparison between A vs B, A vs C and B vs C. I tried with > fit <- lmFit(MA, design=c(1,2,3,-1,-2,-3,1,2,3,-1,-2,-3,1,2,3,-1,-2,-3)) > > but I get results that are too much different for t-test among > conditions, so I think it is wrong. > > thank you > > Claudio Hi Claudio, take a look at the Limma users guide, Chapter 7. It tells you how to create a design and contrasts matrix, etc. Functions like 'model.matrix', 'makeContrasts' and 'contrasts.fit' will make the process quite straightforward. Jose -- Dr. Jose I. de las Heras Email: J.delasHeras at ed.ac.uk The Wellcome Trust Centre for Cell Biology Phone: +44 (0)131 6513374 Institute for Cell & Molecular Biology Fax: +44 (0)131 6507360 Swann Building, Mayfield Road University of Edinburgh Edinburgh EH9 3JR UK