how to normalize data for rna-seq after limma for heatmap or cox or other downstream analysis, should I use the same way as in edger, jusr use the logCPM data here thanks a lot
dge <- DGEList(counts=mRNAdata)
dge <- calcNormFactors(dge)
**logCPM** <- cpm(dge, log=TRUE, prior.count=3)
v <- voom(dge,design,plot=TRUE, normalize="quantile")
fit <- lmFit(v, design)
cont.matrix=makeContrasts(contrasts=c('tumor-normal'),levels = design)
fit2=contrasts.fit(fit,cont.matrix)
fit2=eBayes(fit2)
tempOutput = topTable(fit2, coef='tumor-normal', n=Inf)
DEG_limma_voom = na.omit(tempOutput)
so limma does not have its own method, but can only use the function of edger? can I use the result of voom?
You mean functions for heatmaps and Cox regression? - for Heatmaps, you can use
gplots
,pheatmap
, orComplexHeatmap
( see here: https://github.com/kevinblighe/E-MTAB-6141 ). For Cox regression, you can usesurvival::coxph
For the heatmap, in a typical scenario, you would subset your
logCPM
data-matrix for statistically significant genes identified viatopTable()
, and then perform clustering on this subset data-matrix.no, I am not meaning this I am talking about whether I can use the voom data for downtream analysis instead of logcpm in packages of edger.