Hi,
I am using EdgeR to find DE genes in an experiment with two factors 
(Tissue (2 levels) and Treatment (3 levels). Currently the design matrix 
output by model.matrix() looks like this:

        Int     Tissue  Treat1  Treat2  Treat3 
S1      1       0       0       0       1
S2      1       0       0       0       1
S3      1       0       0       1       0
S4      1       0       0       1       0
S5      1       0       1       0       0
S6      1       0       1       0       0
S7      1       1       0       0       1
S8      1       1       0       0       1
S9      1       1       0       1       0
S10     1       1       0       1       0
S11     1       1       1       0       0
S12     1       1       1       0       0
attr(,"assign")
[1] 0 1 2 2 2
attr(,"contrasts")
attr(,"contrasts")$Tissue
[1] "contr.treatment"
attr(,"contrasts")$Treat
[1] "contr.treatment"

and the model is a comparison of this design and the model without the 
last column.

I would like to specify a design matrix where a) I drop each of the Treat 
levels (in separate runs of EdgeR) to find the DE lists corresponding to 
each level of the Treatment  and to b) specify a model where level1 of the 

Treatment is the reference against which the other levels are compared. 

I am stuck on how to specify the alternate design matrix and how to pass 
this to the model.matrix() command (or even if I have to do this). I am 
aware this is a general R command and have read the manual page and the 
manual pages for the contrasts() function.  My inability to make logical 
sense of these is no doubt testament to my lack of R expertise and some 
help would be appreciated. 

In reading the limma documentation it indicates that a reference can be 
specified by the option ref=" ". Would this also work for me e.g. 
model.matrix(targets, ref="Treat1")?

For specifying the alternate contrasts would this also be achieved in a 
similar way to that shown in the limma manual by setting up a contrasts 
matrix e.g. 
contrast.matrix <- makeContrasts(Treat2,levels=design) to achieve also the 

contrast of Treat2 (still using Treat1 as a reference)?

My next question is then the obvious, how is this then used in the 
testing. Do I specify it in the glmFit() or using the glmLRT() function on 

the original output from the glmFit but specifying the contrasts matrix as 

the option? e.g 
glm1 <- glmFit(DGEList.object, design, dispersion = common.dispersion) # 
fits the model
lrt1 <- glmLRT(DGEList.object, glm1) ## fits the original design contrast
lrt2 <- glmLRT(DGEList.object, glm1, contrast.matrix) ## fits the second 
contrast

or, am I way off track......
looking forward to being set straight.

Josquin

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