# [BioC] edgeR factorial experiment contrasts

James W. MacDonald jmacdon at uw.edu
Wed Sep 3 23:38:24 CEST 2014

```Hi Jahn,

No, it's not wrong. You can parameterize any number of ways, and you will
get identical results for equivalent contrasts. But given that you will get
identical results, and one way is conceptually easier to figure out, I
would in general advocate using the conceptually easier parameterization
over a more difficult one that requires lots of algebraic contortions in
order to figure out what contrast to use.

I believe you sent a truncated version of your experiment (there were no
replicates at each time point), so I can't give you explicit code. However,
say you have a design matrix that looks something like this:

E00 J00 E01 J01 E05 J05 E48 J48 E96 J96
<zeros and ones go here>

And you want to compare the third contrast you mention below. It's as
simple as

glmLRT(fit, contrast = c(0.33, 0, 0.33, 0,0.33, 0,-0.5, 0, -0.5, 0))

Which tests the average of the E0-5 time points against the average of the
E48-96 time points.

And interactions are simple as well. Say you want the interaction between
times 0 and 96:

glmLRT(fit, contrast = c(1,-1,0,0,0,0,0,0,-1,1))

Simple, no?

Best,

Jim

On Wed, Sep 3, 2014 at 4:10 PM, Jahn Davik <jahn.davik at bioforsk.no> wrote:

>  Hi Jim,
> Yes, I believe I can see that (except for possibly my third question).
> It's just that I tend to think along these lines. Are they wrong?
>
> jahn
>
>  ------------------------------
> *Fra:* James W. MacDonald [jmacdon at uw.edu]
> *Sendt:* 3. september 2014 22:06
> *Til:* Jahn Davik [guest]
> *Kopi:* bioconductor at r-project.org; Jahn Davik
> *Emne:* Re: [BioC] 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] <
> guest at bioconductor.org> 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 (BokmÃ¥l)_Norway.1252  LC_CTYPE=Norwegian
>> (BokmÃ¥l)_Norway.1252
>> [3] LC_MONETARY=Norwegian (BokmÃ¥l)_Norway.1252 LC_NUMERIC=C
>> [5] LC_TIME=Norwegian (BokmÃ¥l)_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
>

--
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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