[BioC] Analyze miRNA experiment in Bioconductor

[Paul Geeleher]

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?

-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
>  >
>
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