[BioC] backgroundCorrect offset value
James W. MacDonald
jmacdon at med.umich.edu
Fri Aug 26 22:12:57 CEST 2011
On 8/26/2011 11:00 AM, Prasad Siddavatam wrote:
> Hello Users,
> I have a question regarding the usage of backgroundCorrect function in LIMMA.
> when I do the following with offset 50, I am getting 2900 differentially
> expressed genes
> RG.b<- backgroundCorrect(RG, method = "normexp", offset = 50);
> where as, when I do the following with offset 1,
> I am getting 1300 differentially expressed genes
> RG.b<- backgroundCorrect(RG, method = "normexp", offset = 1);
> Please advise which offset value to be used? Why is offset value making
> so much difference?
I can't advise you on the offset to use; that is up to you as the data
analyst. But I can explain why you get more genes with a larger offset.
When you do a local background correction of your data, for the set of
spots that are fairly dim (not much different from background
intensity), the resulting ratios can become unstable because the
numerators and/or denominators get small. This gives the characteristic
spreading of the MA plot at low intensities after background correction.
An extreme example would be the instance where the R and G channels are
nearly identical (say, 200 and 205), so the uncorrected ratio is close
to 1. But if the Rb and Gb values are, say 190 and 185, then the
background corrected ratio will be 2! Adding 50 to the R and G values
before background correction will dampen the ratio to 0.86, which is
likely closer to truth.
If you do MA plots before background correction and then after, both
with and without adding the offset you will see what I mean.
> I am using this for TWO channel data, which is read by "genepix".
> Greatly appreciate your help.
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James W. MacDonald, M.S.
University of Michigan
Department of Human Genetics
1241 E. Catherine St.
Ann Arbor MI 48109-5618
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