Re: edgeR factorial experiment contrasts

Hi Jahn,

Things will go much easier for you if you just parameterize as cell means
rather than a conventional factorial model.

grps <- factor(paste(Group, Time, sep = "_"))

design <- model.matrix (~0+grps)

Then all the comparisons are easy to do.

Best,

Jim



On Wed, Sep 3, 2014 at 12:39 PM, Jahn Davik [guest] <[email protected]>
wrote:

> Hi,
> I am trying to figure out how to do comparisons using edgeR but have to
> admit that I am on unsure ice.
> My experiment is abiotic stress on two genotypes (E and J) and these have
> been sampled for RNA-seq at five time points (00, 01, 05, 48, and 96).
> I would like to analyze this experiment as a factorial (which I am sure
> could be argued), and when I set up the following design:
>
> > design<-model.matrix(~0 + Group + Time + Group*Time),
>
> I get the following design matrix (I've taken out the 3 three replicates
> at experimental cell.):
>
> > design
>       GroupE GroupJ Time01 Time05 Time48 Time96 GroupJ:Time01
> GroupJ:Time05 GroupJ:Time48 GroupJ:Time96
> E00      1      0      0      0      0      0             0             0
>            0             0
> J00      0      1      0      0      0      0             0             0
>            0             0
> E01      1      0      1      0      0      0             0             0
>            0             0
> J01      0      1      1      0      0      0             1             0
>            0             0
> E05      1      0      0      1      0      0             0             0
>            0             0
> J05      0      1      0      1      0      0             0             1
>            0             0
> E48      1      0      0      0      1      0             0             0
>            0             0
> J48      0      1      0      0      1      0             0             0
>            1             0
> E96      1      0      0      0      0      1             0             0
>            0             0
> J96      0      1      0      0      0      1             0             0
>            0             1
> attr(,"assign")
>  [1] 1 1 2 2 2 2 3 3 3 3
> attr(,"contrasts")
> attr(,"contrasts")$Group
> [1] "contr.treatment"
>
> attr(,"contrasts")$Time
> [1] "contr.treatment"
>
>
> I have some questions regarding testing contrasts:
> First though, I realize that I can pull out the data for E00 and J00, say,
> and compare these using exactTest, but then I guess that you are
> disregarding all information that lies in the remaining data set? That does
> not seem to me to be a good way to do it. But I may be wrong here?
>
> Second, if I want to do the comparison E00 vs J00 using the design
> indicated above, how should that be set up? Would
>
> lrt <- glmLRT(fit,contrast=c(1,-1,0,0,0,0,0,0,0,0)) do it?
>
> Third, if I want to compare (E00 + E01 + E05) vs (E48 + E96), how would I
> set up that contrast?
> For my own pedagogic reasons I can show how I was thinking in this case:
>
> E00 = L1
> E01 = L1 + L4   x 2  -->  6L1 + 2L3 + 2L4
> E05 = L1 + L4
>
> versus                                       --> 2L3 + 2L4 - 3L5 -3L6
>
> E48 = L1 + L5   x 3  -->  6L1 + 3L5 + 3L6
> E96 = L1 + L6
>
>
> resulting in the contrast:
>
> lrt <- glmLRT(fit,contrast=c(0,0,2,2,-3,-3,0,0,0,0))
>
> Then I seem to remember that contrasts should sum up to zero? If anybody
> could tell me where I am loosing it, I would be very obliged.
>
> Thank you
>
> jd
>
>
>
>
>
>  -- output of sessionInfo():
>
> R version 3.1.0 (2014-04-10)
> Platform: x86_64-w64-mingw32/x64 (64-bit)
>
> locale:
> [1] LC_COLLATE=Norwegian (Bokmål)_Norway.1252  LC_CTYPE=Norwegian
> (Bokmål)_Norway.1252
> [3] LC_MONETARY=Norwegian (Bokmål)_Norway.1252 LC_NUMERIC=C
> [5] LC_TIME=Norwegian (Bokmål)_Norway.1252
>
> attached base packages:
> [1] splines   stats     graphics  grDevices utils     datasets  methods
>  base
>
> other attached packages:
> [1] edgeR_3.6.8          limma_3.20.9         BiocInstaller_1.14.2
>
> loaded via a namespace (and not attached):
> [1] tools_3.1.0
> >
>
> --
> Sent via the guest posting facility at bioconductor.org.
>
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-- 
James W. MacDonald, M.S.
Biostatistician
University of Washington
Environmental and Occupational Health Sciences
4225 Roosevelt Way NE, # 100
Seattle WA 98105-6099

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