Hello Upendra

I have a similar question about setting up the model matrix in edgeR for performing anova-like analysis. Since I am new to computing in general, I don't entirely understand the codes.

Experimental setup: I have two mammalian cell lines (Lets say) A, B. RNA-Seq analysis was done on these two cell lines at different concentrations of serum: 10%, 5% and 0%. Only cell line A was present in all three conditions (10,5,0) but cell line B is present in only 10% and 5%.

I want to do an anova analysis across the two cell lines in all conditions. Could you please verify the way I have setup the model matrix? My code is below.

##Creating the DGE List

> group<-c(rep("P_10",2), rep("P_05",2), rep("P_00",2), rep("H_10",2), rep("H_05",2))
> cds<-DGEList(counts, group=group)

# Calculating Normalization factors
> cds<-calcNormFactors(cds)

#MDS Plot
> plotMDS(cds,main='MDS plot for count data', labels=colnames(cds$counts))
> cds<-estimateCommonDisp(cds)
> cds<-estimateTagwiseDisp(cds,prior.df = 10)

#model matrix code
> design<-model.matrix(~0+group, data=cds$samples)

#Anova like analysis code
> cds<-estimateGLMCommonDisp(cds,design)
> cds<-estimateGLMTrendedDisp(cds,design)
Loading required package: splines

> cds<-estimateGLMTagwiseDisp(cds,design)
> fit2<-glmFit(cds,design)
> lrt.anova_like<-glmLRT(fit2,coef=2:5)

Thank You