Is limma suitable for identifying differentially expressed genes across species?
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Entering edit mode
6.3 years ago
EvoGen ▴ 10

For microarray gene expression data for

5 species : sp1,sp2,sp3 ...

7  time-points : t1,t2,t3...

3 replicates: r1,r2,r3 ...

And the aim is to identify genes differentially expressed genes w.r.t to sp1(assume sp1 is reference species with whom i want compare expression values).

Using limma i compared created a contrast matrix to compare sp2-sp1,sp3-sp1,sp3-sp1 .

design matrix looks like :

design <- model.matrix(~0+ targets$Organism)
colnames(design) <- c("sp1","sp2","sp3","sp4","sp5","sp6")
> head(design)
  sp1 sp2 sp3 sp4 sp5 sp6
1   0   1   0   0   0   0
2   0   1   0   0   0   0
3   0   1   0   0   0   0
4   0   1   0   0   0   0
5   0   1   0   0   0   0
6   0   1   0   0   0   0
> tail(design)
    sp1 sp2 sp3 sp4 sp5 sp6
139   0   0   0   0   0   1
140   0   0   0   0   0   1
141   0   0   0   0   0   1
142   0   0   0   0   0   1
143   0   0   0   0   0   1
144   0   0   0   0   0   1

> contrast.matrix <- makeContrasts(sp2-sp1,sp3-sp1,sp4-sp1,sp5-sp1,sp6-sp1,levels=design)
> contrast.matrix
      Contrasts
Levels sp2 - sp1 sp3 - sp1 sp4 - sp1 sp5 - sp1 sp6 - sp1
   sp1        -1        -1        -1        -1        -1
   sp2         1         0         0         0         0
   sp3         0         1         0         0         0
   sp4         0         0         1         0         0
   sp5         0         0         0         1         0
   sp6         0         0         0         0         1

But for e.g for sp2-sp1 set , not only genes differentially expressed in sp2 but also differentially expressed in some other species e.g sp4 is given as output.

 

So, i am not sure if limma package is suitable for such time-point and species specific data ?

Can anyone suggest any other approach or package to identify genes differentially expressed across species ?

 

limma microarray timecourse gene expression • 1.8k views
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will limma work for samples without replicates?

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6.3 years ago

Limma is a completely appropriate tool for this sort of experiment (in fact, it'd probably be the most commonly used tool). Just because a particular gene is differentially expressed between sp2 and sp1 doesn't mean it can't also be DE between sp4 and sp1 (or even between sp4 and sp2).

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ok.. i completely agree with you that a gene can be DE in multiple species .

But would there be any means to identify or pin-point genes whose expression is conserved in sp1,sp3,sp4 but DE only for sp2 .?

What if i group sp1,sp2,sp4 together say into allsps and compare sp2-allsps , to get DE in sp2 against all the remaining species ?

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For that sort of thing it's often best to use something like WGCNA. I say "better" because I suspect there are a bunch of patterns like this that you'd like to look at and this is a convenient* method to do that.

*This does not imply that WGCNA is easy to use and/or understand/debug.

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