Question: DESeq2: before differential analysis
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gravatar for limin201709
10 months ago by
limin20170910
limin20170910 wrote:

I was doing RNA seq data process, and I would like to normalize the raw count and +0.1 to avoid the zero log2 fold change. But if I +1 to normalize counts data, it becomes a matric and how can I continue the pipeline.

library(DESeq2)
L3Ddata = FG_20141202_TF_20160315_aspFumA1163_RNA_seq[,13:18]
L3Ddata<-as.data.frame(L3Ddata)
colnames(L3Ddata)=test$samplename# rows - genes, columns - samples
L3Dcolname=test[,2:4]
rownames(L3Dcolname)=test$samplename##adapt the name 

L3D_dds = DESeqDataSetFromMatrix(L3Ddata,colData = L3Dcolname, design=~condition)
all(rownames(L3Dcolname)%in%colnames(L3Ddata))##test if the colname of count data equals to rowname of description
L3D_dds<-counts(L3D_dds,normalize=TRUE)
L3D_dds=L3D_dds+0.1  #here it becomes a matrix
#normalize counts +0.1 to avid zero counts
L3D_dds<- DESeq(L3D_dds)# takes quite some time
rna-seq R • 324 views
ADD COMMENTlink modified 10 months ago by Devon Ryan86k • written 10 months ago by limin20170910
0
gravatar for Devon Ryan
10 months ago by
Devon Ryan86k
Freiburg, Germany
Devon Ryan86k wrote:

There is absolutely no reason to add a pseudocount to your actual counts. A log2 fold change of 0 occurs when there's no change and has nothing to do with there being 0s in the data. If you're worried about extreme fold-changes due to having zeros, then that's what the lfcShrink() command is for.

ADD COMMENTlink written 10 months ago by Devon Ryan86k

sorry, it is to avoid zero counts.

ADD REPLYlink written 10 months ago by limin20170910
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