[BioC] Background correction - limma

Gordon Smyth smyth at wehi.edu.au
Wed Apr 7 01:32:22 CEST 2004

At 10:29 PM 6/04/2004, Helen Cattan wrote:
>I am trying to use backgroundCorrect - method = half or minimum but get
>the following error:
> > files=dir(pattern="*\\.gpr")
> > RG=read.maimages(files, columns=list(Rf="F635 Median", Gf="F532
>Median", Rb="B635 Median", Gb="B532 Median"))
> > names(RG)
> > RG$genes=readGAL()
> > RG$printer=getLayout(RG$genes)
> > samples=read.table("sampleinformationa.txt", header=TRUE, sep="\t",
> > samples
> > spottypes=readSpotTypes()
> > RG$genes$Status=controlStatus(spottypes, RG)
> > RG = backgroundCorrect(RG, method="minimum")
> > MA=normalizeWithinArrays(RG, RG$printer)
> > MA=normalizeBetweenArrays(MA)
> > design=c(1,-1)
> > cor=dupcor.series(MA$M,design,ndups=2)
>Loading required package: nlme
>Loading required package: lattice
>Error in array(val[["varBeta"]], c(rnkm1, rnkm1), list(namCoef,
>namCoef)) :
>         length of dimnames[1] not equal to array extent

The error seems to be occuring in the function 'gls' from the nlme library 
which is called by dupcor.series(). Congratulations, you're the first 
person I know of who has managed to break gls()!

Can I ask you to upgrade to the current version of limma from 
http://bioinf.wehi.edu.au/limma and try again. dupcor.series() no longer 
calls gls().

If you still have a problem, please type summary(MA$M) to see how many 
missing values you have in your data.

>Then I manually altered the background values of the genepix files so
>after background subtraction the previously negative (F635/532-B635/532)
>values were zero,

Altering negative values to zero will make no difference. log of zero is 
negative infinity. Such values will be removed by dupcor.series() and other 
regression programs just as if they were missing values. If you want to do 
something comparable to method="minimum" you have to replace with a 
positive value.


>  omitted the backroundCorrect method and this worked
>fine. However when I redid the alteration so after background
>subtraction the previously negative values were 0.5, I got the same
>error as above.
>Has anyone any suggestions?

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