[BioC] LIMMA: MA, design, and contrasts

[Tiandao Li]

Hello,

I am using limma for 2-color microarray data analysis. I have some 
questions regarding MA, design, and contrasts.

1. I have one MA file including all my experiments. Rows of MA correspond 
to spots and columns to individual experimental file. Columns were listed 
as the increasing order of file names, since I used barcodes as file 
names. Then after reading in the target file and creating design matrix, I 
used the following to caluculate correlation between duplicates.

corfit <- duplicateCorrelation(MA,design,ndups=4) # A slow computation!
corfit$consensus.correlation

However if columns of MA2 are listed following the order of 
target$FileName, corfit2$consensus.correlation value is different from one 
of MA. Which one should I use in the next anlysis?

2. I have one MA file including all my experiments, and also an all-in-one 
contrast matrix including different contrasts (related or un-related). 
Should I use this all-in-one contrast matrix for linear model to find the 
differentially expressed genes? This doesn't sound right. Or I use subset 
of MA for and only for related one or more contrasts, and use all-in-one 
MA only necessary. Which one is better?

Thanks in advance,

Tiandao