edgeR: design matrix for methylation analysis
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sgld • 0
@sgld-16356
Last seen 3.0 years ago

hi,

when I using edgeR to identify differentially methylated regions (DMR) between different groups with edgeR guidepaper, I have a question about the design matrix .

######

> sam <- rep(samples, each=2)
> meth <- factor(rep(c("Me","Un"),6), levels=c("Un","Me"))
> design <- model.matrix(~ sam + meth)
> colnames(design) <- gsub("sam","",colnames(design))
> colnames(design) <- gsub("meth","",colnames(design))
> colnames(design)[1] <- "Int"
> design <- cbind(design,
+ Me2=c(0,0,0,0,1,0,1,0,0,0,0,0),
+ Me3=c(0,0,0,0,0,0,0,0,1,0,1,0))

 The 7th column “Me” represents the methy-lation level (or M-value) in the 40-45 µm group. The 8th column “Me2” represents the differencein methylation level between the 50-55 and the 40-45 µm groups. Finally, the last column “Me3”
represents the difference in methylation level between the 60-65 and the 40-45 µm groups.

######

the contents above from edgeR guide, my question is how to to design the contrast column like 8th column.

 

 

edger methylation • 622 views
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intercept group is 40-45cm?

if i want to compare the 50-55cm group to 60-65cm group,how to do?I get a answer ,design matrix in GLM .is it ok for edgeR?

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the method of edgeR or limma to identify differential  analysis is that regressing each group with linear model,then contrast the different groups' regression to identify difference ? 

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@gordon-smyth
Last seen 8 hours ago
WEHI, Melbourne, Australia

You are probably following the edgeR methylation workflow from F1000R:

https://f1000research.com/articles/6-2055/

Since publishing that paper, we have added a new function modelMatrixMeth to edgeR to make it much easier for you. You can now simply use

design <- modelMatrixMeth(~group)

where group is your sample grouping factor. Have a look at the revised methylation workflow, which is here:

http://www.statsci.org/smyth/pubs/edgeRMethylationPreprint.pdf

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thank you for your kindly reply and the nice paper.

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Gordon,what is the role of library size in the differential methylation analysis.  I found there are many "keep.lib.sizes=Fasle " to causes the library sizes to be recomputed in the F1000Research paper.

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The article says:

"In the above code, the two library sizes for each sample should be equal. Otherwise, the library size values
are arbitrary and any settings would lead to the same P-value."

Hence the library sizes must not be recomputed automatically, otherwise they would no longer be equal for the two counts from the same sample.

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I'm very interested with the updated methylation workflow but the link is broken. Can someone provide the link to that document?

Thanks

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The statsci.org website was temporarily down. It's up again now.

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