limma and ebayes : error message : differences in versions?
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Sue Jones ▴ 40
@sue-jones-1758
Last seen 9.6 years ago
I have used limma to calculate differential gene expression (see code at end of message). I am currently using R 2.7, Biobase 2.0.1 and Limma 2.14.6 and get the error message Warning message: Zero sample variances detected, have been offset after uisng eBayes. I know a previous message to this list explains this and says it can be ignored. However, I conducted the same analysis using older versions of R, Limma and Biobase and did not see this message and got a much larger list of differentially expressed genes. Using the current versions of limma and biobase I see this message and my list of differentially expressed genes is smaller - can this be correct? Thanks for anyones help. Code ------------------------------------------------------ > files <- c("W05h_1.CEL", "W0h_1.CEL", "W2h_1.CEL", + "W2h_2.CEL", "W4h_1.CEL", "GW_025.CEL", "GW_1.CEL", "GW_3.CEL","E2h_1.CEL", "E2h_2.CEL", + "E2h_3.CEL", "E2h_4.CEL") > Data <- ReadAffy(filenames = files) > Data_gcrma <- gcrma(Data) > design<- cbind(WAT=c(1,1,1,1,1,1,1,1,0,0,0,0), ETH=c(0,0,0,0,0,0,0,0,1,1,1,1)) > fit <- lmFit(Data_gcrma,design) > cont.matrix <- makeContrasts(WATvsETH=ETH-WAT, levels=design) > fit2<- contrasts.fit(fit,cont.matrix) > fit3 <- eBayes(fit2) Warning message: Zero sample variances detected, have been offset Sue Jones University of Sussex
Biobase limma Biobase limma • 3.4k views
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@james-w-macdonald-5106
Last seen 8 hours ago
United States
Hi Sue, Sue Jones wrote: > > > I have used limma to calculate differential gene expression (see code at > end of message). I am currently using R 2.7, Biobase 2.0.1 and Limma > 2.14.6 and get the error message > > Warning message: > Zero sample variances detected, have been offset That's not an error - it's a warning. The difference being that your analysis proceeded, but there was something you might want to know about versus your computer going crash bang thud and halting. > > after uisng eBayes. > > I know a previous message to this list explains this and says it can be > ignored. > > However, I conducted the same analysis using older versions of R, Limma > and Biobase and did not see this message and got a much larger list of > differentially expressed genes. Using the current versions of limma and > biobase I see this message and my list of differentially expressed genes > is smaller - can this be correct? Yes, but probably for a reason different from the one you mention. You are using gcrma() to generate expression values and there have been several comments on this list about more recent versions of gcrma() generating summary statistics that are identical for all samples (hence zero sample variance). So, is this a problem? I don't know. It is possible that your previous list of differentially expressed genes had more false positives than your current list. Or not. That's the problem with much of this high throughput data - you get tons of data, but there is no ground truth so it is difficult to separate the signal from the noise. Best, Jim > > Thanks for anyones help. > > > Code > ------------------------------------------------------ >> files <- c("W05h_1.CEL", "W0h_1.CEL", "W2h_1.CEL", > + "W2h_2.CEL", "W4h_1.CEL", "GW_025.CEL", "GW_1.CEL", > "GW_3.CEL","E2h_1.CEL", "E2h_2.CEL", > + "E2h_3.CEL", "E2h_4.CEL") >> Data <- ReadAffy(filenames = files) >> Data_gcrma <- gcrma(Data) >> design<- cbind(WAT=c(1,1,1,1,1,1,1,1,0,0,0,0), >> ETH=c(0,0,0,0,0,0,0,0,1,1,1,1)) >> fit <- lmFit(Data_gcrma,design) >> cont.matrix <- makeContrasts(WATvsETH=ETH-WAT, levels=design) >> fit2<- contrasts.fit(fit,cont.matrix) >> fit3 <- eBayes(fit2) > Warning message: > Zero sample variances detected, have been offset > > > Sue Jones > University of Sussex > > _______________________________________________ > Bioconductor mailing list > Bioconductor at stat.math.ethz.ch > https://stat.ethz.ch/mailman/listinfo/bioconductor > Search the archives: > http://news.gmane.org/gmane.science.biology.informatics.conductor -- James W. MacDonald, M.S. Biostatistician Douglas Lab University of Michigan Department of Human Genetics 5912 Buhl 1241 E. Catherine St. Ann Arbor MI 48109-5618 734-615-7826
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