[Bioc-devel] Limma 2.13.7: VSN normalization

[Koen Van Leemput]

Hello,

In preparation of an upgrade of R on our server, I have been testing our 
microarray analysis workflow (making extensive use of the limma and vsn 
packages)  and comparing results between the recent versions I am using 
now (R 2.6.2, limma 2.13.7 and vs 3.4.13)  and the old versions that are 
currently in use (R 2.4.1, limma 2.9.13, vsn 1.12.0).

I have observed differences in the output of the 
"normalizeBetweenArrays" function when using the "vsn" normalization 
method. 

After looking through the documentation of both limma and vsn I think 
the problem lies in the fact that the main normalization function in the 
recent vsn package (called "vsn2") outputs data on a glog scale to base 
2 while the previous version of this function "vsn" outputs data on glog 
scale of the natural logarithm. 

I have looked at the actual call made by limma in version 2.9.13:

       y <- vsn(intensities = y, ...)
        n2 <- ncol(y at exprs)/2
        G <- y at exprs[, 1:n2]/log(2)
        R <- y at exprs[, n2 + (1:n2)]/log(2)


and compared it to the call made in the newer version 2.13.7:

        y <- vsnMatrix(x = y, ...)
        n2 <- ncol(y at hx)/2
        G <- y at hx[, 1:n2]/log(2)
        R <- y at hx[, n2 + (1:n2)]/log(2)

Am I right in thinking that the division by log(2) is not necessary 
anymore in this last case?

As far as I have tested, calling vsn like this:

       y <- vsnMatrix(x = y, ...)
       n2 <- ncol(y at hx)/2
       G <- y at hx[, 1:n2]
       R <- y at hx[, n2 + (1:n2)]

produces very similar results to the old situation.  Is this something 
that should be changed in the new limma version or are the new results 
actually the more correct?

Thanks,
kind regards,
Koen

-- 
Koen Van Leemput
Advanced Database Research and Modelling (ADReM)
Department of Mathematics and Computer Science
University of Antwerp - Campus Middelheim
Middelheimlaan 1, office G.217, B-2020 Antwerpen, Belgium
e-mail: koen.vanleemput at ua.ac.be
tel.: +32 3 265 3407
fax: +32 3 265 3204