Hi everyone, I am new to microarray data analysis. Please evaluate the following code for block analysis in Limma and let me know about whether this code is appropriate??. library(limma) eset<-read.table("RMANormData_apap.txt") design <- model.matrix(~ -1+factor(c(1,1,1,2,2,2,3,3,3,4,4,4,5,5,5,6,6,6))) colnames(design) <- c("control3", "control24", "low3","high3","low24","high24") contrast.matrix <- makeContrasts(high3-control3,high24-control24,low3-control3,low24-cont rol24 , levels=design) sample<-read.table("sample.txt",sep="\t",as.is=T,h=T) corfit<-duplicateCorrelation(eset,design,ndups=1,block=sample[,3]) fit<-lmFit(eset,design,block=sample[,3],cor=corfit$consensus) fit2<-contrasts.fit(fit,contrast.matrix) fit2<-eBayes(fit2) topTable(fit2, coef=1, adjust="fdr", sort.by="B", number=10) results <- decideTests(fit2,adjust.method="BH",method="separate",lfc=1, p.value=0.05); The input table RMANormData_apap.txt containing the expression values of the genes includes the following columns: c_3_1 c_3_2 c_3_3 c_24_1 c_24_2 c_24_3 t_1l_3h_1 t_1l_3h_2 t_1l_3h_3 t_5l_3h_1 t_5l_3h_2 t_5l_3h_3 t_1l_24h_1 t_1l_24h_2 t_1l_24h_3 t_5l_24h_1 t_5l_24h_2 t_5l_24h_3 the input file sample.txt contains the following information: Sample Group Block c_3_1 control3 1 c_3_2 control3 2 c_3_3 control3 3 c_24_1 control24 1 c_24_2 control24 2 c_24_3 control24 3 t_1l_3h_1 low3 1 t_1l_3h_2 low3 2 t_1l_3h_3 low3 3 t_5l_3h_1 high3 1 t_5l_3h_2 high3 2 t_5l_3h_3 high3 3 t_1l_24h_1 low24 1 t_1l_24h_2 low24 2 t_1l_24h_3 low24 3 t_5l_24h_1 high24 1 t_5l_24h_2 high24 2 t_5l_24h_3 high24 3 since the experiment is performed using 3 rats block factor in included. Regards, Sri [[alternative HTML version deleted]]