[BioC] limma package

Gordon Smyth smyth at wehi.edu.au
Thu Dec 11 01:44:56 MET 2003

At 06:15 AM 11/12/2003, Anna Cao wrote:
>I'm trying to normalize between arrays using the limmma package so I
>can run SAM. I want to know whether we can read log2 ratio data into
>R? I'm using Agilent's extraction program to extract cDNA arrays
>data and the image analysis program from Agilent automatically
>normalize within array, giving the normalized intensity and log
>ratio. Is there anyway I can by-pass reading Mean/Median foreground
>and background intensity and directly feed the processed (background
>subtracted and normalized) intensity into R?

This is easy in R. Use the Agilent flat file, use read.table(), then 
collect the normalized columns into a matrix in R. The matrix can then but 
used as input for example to the lmFit() command in limma. You'll have to 
do the input yourself though, there aren't any canned limma commands to do 
it. If you need help with this, you might want to ask for help on the 
R-help mailing list.

>Another question: Can I also input a column that filters spots which
>were unsatisfactory according to the image analysis program? And how
>can I can remove these bad spots from further calculation in R using
>the limma package or any other functions in R?

Using limma, you set the appropriate elements of the 'weights' matrix to 
zero for those spots. This applies to both normalization and linear 
modelling functions in limma. Normally the weights matrix is set 
automatically using the wt.fun argument of read.maimages, but you will need 
to create it yourself if you're reading in the data manually as it were.



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