I have gene expression measured in two biopsies each from 125 animals. One biopsy was untreated, while the other was pre-treated with a drug. The investigator hybridized untreated biopsies to the green channel of an Agilent microarray and treated biopsies to the corresponding red channels. The animals are genetically distinct, so green does not represent a uniform reference.
To get the red/green ratio, I'm using limma:
raw = read.maimages(source="agilent.median", files=file.list, wt.fun=wtAgilent) norm = normalizeWithinArrays(raw, method="loess", bc.method="normexp", offset=50)
I would also like to normalize the red and green channels separately with normalizeBetweenArrays(). I can apparently extract the green channel only with the green.only parameter of read.maimages() , but there is no red.only parameter. What is the best (most elegant and correct) way to extract the separate channels and normalize them? I have a feeling the method="Rquantile" parameter of normalizeBetweenArrays() isn't quite what I want, but I'm not sure what is.