edgeR design: pool groups

[sylvian]

Hi,

I am using edgeR post kallisto to look for DGE.

I have RNAseq samples from 4 different tissue types and my design currently looks like this:

> head(design)
(Intercept) D.new$samples[, 1]CA Endothelium
1 1 1
2 1 1
3 1 1
4 1 1
5 1 1
6 1 1
D.new$samples[, 1]CA Smooth Muscle D.new$samples[, 1]Coronary Artery
1 0 0
2 0 0
3 0 0
4 0 0
5 0 0
6 0 0
...

attr(,"assign")
[1] 0 1 1 1
attr(,"contrasts")
attr(,"contrasts")$`D.new$samples[, 1]`
[1] "contr.treatment"


So my four tissue types are Aorta (intercept), CA Endothelium, CA Smooth Muscle and Coronary Artery.

As I understand I can compare gene expression between any tissue using:
lrt.ECvsAo<-glmLRT(fit.new, contrast=c(1,-1,0,0))


If I use this contrast
lrt.ECvsAll<-glmLRT(fit.new, contrast=c(1,-1,1,1))
am I asking for DGE between the second column (CA Endothelium) and EITHER other tissue?

I'd like to find genes that are differentially expressed between CA Endothelium and ALL Aorta, CA Smooth Muscle and Coronary Artery.

Is this doable?

Many thanks

[lpachter]

If you're quantifying with kallisto you may want to take a look at sleuth

[TRA]

Hi all,
I am using edgeR flow to get gene lists of multifactorial experiment. I have 6 conditions with 3 samples per each. The problem is some of the gene lists I got end at FDR much less than what I entered and so I did not get the whole gene list. For example, the upregulated genes were completely missing in some lists. I tried increasing the number of genes the program gives using the
< gene_table<-topTags(lrt, n=40000)
but did not help. Then I tried very restreignent FDR as follow
< DE<-tp$table[tp$table$FDR<0.000001,]$GeneID
But again did not help. The last gene in one of my lists was with
Symbol Chr logFC logCPM LR PValue FDR
Hpx 7 -7.186931303 13.83923493 57.6010426 3.21E-014 3.23E-014

Any help would be appreciated!