Confused about using ComBat_seq with voom and limma? Should run batch as a covariate and use uncorrected instead?
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Ahdee ▴ 50
@ahdee-8938
Last seen 6 weeks ago
United States

Hi my understanding is that we should not use corrected counts with something like edgeR or limma?

However, because ComBat_seq is correcting at the raw count level and then input into edgeR for TMM follow with voom can I use this for downstream limma or should I just run the voom limma with uncorrected and add the batch as covariate?

A

sva limma • 154 views
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@kevin
Last seen 1 hour ago
Republic of Ireland

should I just run the voom limma with uncorrected and add the batch as covariate?

This is the preferable option. Also, by including batch as a covariate in a model with formula of, e.g., ~ condition + batch, the test statistics that you derive for condition will be adjusted for the effects of batch.

Following this approach, if you later wish to obtain batch-corrected expression levels for downstream purposes like heatmap + clustering, classification algorithms, et cetera, then use limma::removeBatchEffect() on the output of cpm(y, log = TRUE)

A related question and answers: edgeR and removeBatchEffect()

Kevin

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thanks. This is what i thought as well but the correction at the raw count level got me second guessing.

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