[BioC] Limma lmFit function and spot quality weights

[Benoit]

Hello,
I'm using Limma to assess differential expression on double colour 
microarray data and have a question about the lmFit function.
When I fit linear model using lmFit, as I understood, the function uses 
the weights extracted from the MA object when present and/or specified.
Thus, I tried fitting with and without the spot quality weights and I 
found different results (not very surprising in fact).
In fact, when I used weights, zero weighted spots seemed to be removed 
from the analysis and it's here that I have a problem.

For my experiment, I compare two groups (control vs treated) in a 
classical design experiment "Two Groups: Common Reference" as describe 
in the Limma documentation.

design=modelMatrix(targets,ref="ref")
design
fit=lmFit(MA,design,weights=MA$weights)
/alternative without weights : fit=lmFit(MA,design,weights=NULL)/
cont.matrix=makeContrasts(pollutedVScontrol=polluted-control,polluted,control,levels=design)
cont.matrix
fit2=contrasts.fit(fit,cont.matrix)
fit2=eBayes(fit2)
res=toptable(coef=1,number=15744,fit=fit2,genelist=fit2$genes,adjust.method="BH",A=fit2$Amean,eb=fit2,p.value=0.01)

The difference between the analysis with and without weights is that 
when I use weights new genes highly differentially expressed appeared.
When I control these genes, in fact they correspond to spots that are 
flagged (0) on the majority of the arrays (i.e. only one weight at 1 for 
the control and one weight at 1 for the treated). Thus for these genes 
the comparison is performed only one "control array" versus one "treated 
array".
So is it possible to specify to lmFit that there must be a minimum of 
"1" weights or a maximum "0" weights per groups of array ?

Thank you for any help you can bring me.

Benoit

-- 
Benoit Loup, PhD
UMR Biologie du Développement et Reproduction
Différenciation des Gonades et Perturbations
INRA – Domaine de Vilvert
Bâtiment Jacques Poly
78350 Jouy en Josas
France

Tel: 33 1 34 65 25 38
Fax: 33 1 34 65 22 41
E-mail: benoit.loup at jouy.inra.fr