Doesn't seem to be anything in the users guide specific to this kind of analysis unfortunately. -Paul On Thu, May 8, 2008 at 10:31 AM, Wolfgang Huber <huber at="" ebi.ac.uk=""> wrote: > Dear Paul, > >> Hmm interesting. I might try introducing the extra columns into the >> files and specifying all the values as 0. I can't see why that >> shouldn't work? > > It might, but Narendra's suggestion of reading the limma users guide is a > worthwhile option to consider. > > Best wishes > Wolfgang > > ------------------------------------------------------------------ > Wolfgang Huber EBI/EMBL Cambridge UK http://www.ebi.ac.uk/huber > >> >> -Paul >> >> On Wed, May 7, 2008 at 1:39 PM, Narendra Kaushik >> <kaushiknk at="" cardiff.ac.uk=""> wrote: >>> >>> You can specify your red channel like this: >>> >>> RG <- read.maimages(files,source="genepix", columns=list(R="F635 >>> Median",G="F532 >>> Median",Rb="B635",Gb="B532")) >>> >>> I will suggest you read limma guide. >>> >>> But I think your have data from Imagene package which gives one file for >>> each channel, you can: >>> >>> files <- targets[,c("FileNameCy3","FileNameCy5")] >>> RG <- read.maimages(files, source="imagene") >>> >>> Hope, this helps >>> >>> Narendra >>> >>> >>> "Paul Geeleher" <paulgeeleher at="" gmail.com=""> 07/05/2008 13:24:01 >>> >>> >>> >>> Hi Deepayan, >>> >>> Thanks for your reply. I suppose my main concern is how I should read >>> in the data initially in order to be able to use the normal tools to >>> analyze the data. Reading the data normally like this: >>> >>> RG <- read.maimages( files, source="genepix") >>> >>> Gives the following error: >>> >>> Error in RG[[a]][, i] <- obj[, columns[[a]]] : >>> number of items to replace is not a multiple of replacement length >>> >>> >>> I'm assuming this is down to the fact that the files only contain >>> intensity data for one color rather than two? >>> >>> How should I go about reading the data? >>> >>> Thanks alot, >>> >>> -Paul. >>> >>> On Tue, May 6, 2008 at 10:15 PM, Deepayan Sarkar >>> <deepayan.sarkar at="" gmail.com=""> wrote: >>> > On 5/6/08, Paul Geeleher <paulgeeleher at="" gmail.com=""> wrote: >>> > > Dear Members, >>> > > >>> > > I've inherited a bunch of GenePix files from an miRNA experiment. >>> They >>> > > are single color arrays, ( as opposed to 2 color as is the norm >>> for >>> > > GenePix I think). There is a subset of 7 arrays and I wish to >>> compare >>> > > a group of 4 of these to the other group of 3 and analyze >>> differential >>> > > expression between the two groups. I was hoping somebody could >>> point >>> > > me in the right direction of how I'd go about doing this with >>> > > Bioconductor? Is it possible using the Limma package? Is there any >>> > > code out there to assist me? >>> > > >>> > > I've experience in analyzing Affymetrix data using Limma and PUMA, >>> but >>> > > not GenePix, and the Limma Users Guide seems to focus on analyzing >>> two >>> > > dye experiments. >>> > >>> > Any analysis ultimately boils down to some sort of normalization, and >>> > the actual differential expression analysis. The second part in limma >>> > (lmFit, etc.) can work with any expression matrix, irrespective of >>> > whether it's 2-color or 1-color (or affy). >>> > >>> > We have been working with a miRNA array dataset recently, and we used >>> > limma to read in the GPR files and do the differential expression >>> > analysis (on one channel). For normalization, many of the standard >>> > microarray algorithms probably don't make much sense, but VSN seems >>> to >>> > work fine. >>> > >>> > We don't really have code (beyond what's already in limma and vsn) >>> > that is generally useful; most of the work is in figuring out which >>> > rows are of interest (i.e., those representing human miRNAs), >>> > combining the replicates (you seem to have four of each), etc. I'm >>> > happy to give you more details if you are interested. >>> > >>> > -Deepayan >>> > >