On Mon, Apr 27, 2009 at 11:25 AM, Paolo Innocenti <paolo.innocenti@ebc.uu.se
> wrote:

> Hi all,
>
> I have dataset of 120 Affy arrays, 60 males and 60 females.
> The expression profiles of the 2 groups differs dramatically, i.e. if I run
> a standard RMA + limma, I have ~90% of the genes differentially expressed.


You'll probably want to provide the code that you used to perform the RMA
and limma analyses.  It is difficult to comment on the results of an
analysis without seeing what was done to produce them.  Also, be sure to
provide sessionInfo() in case there are questions about what packages are
used.

Sean



> Also, downregulated genes are twice as many than upregulated genes,
> although if I impose a cutoff of two-fold difference in expression, they are
> almost equal (15% up and 15% down).
> This is clearly breaking the assumption that most of the genes on the array
> should not be differentially expressed, but the result is in line with the
> current knowledge of sex-biased gene expression in my model organism.
>
> I have done some quality control plots, available here:
> - Boxplot:
> http://www.iee.uu.se/zooekol/pdf/hemiarray_qc_boxplot.pdf
>
> - Frequency histogram:
> http://www.iee.uu.se/zooekol/pdf/hemiarray_qc_histogram.pdf
>
> - RLE and NUSE plots:
> http://www.iee.uu.se/zooekol/pdf/hemiarray_qc_RLEandNUSE1.pdf
>
> - CorrelationPlot:
> http://www.iee.uu.se/zooekol/pdf/hemiarray_qc_correlationplot.pdf
>
> - PCA, after RMA normalization:
> http://www.iee.uu.se/zooekol/pdf/hemiarray_qc_pca.pdf
>
> Now, my questions are:
> 1) Is my issue really a issue? If so, how can I perform a robust
> normalization of my arrays?
>
> 2) Is there a tool to assess how "robust" your pre-processing method is in
> respect to this issue?
>
> 3) Sex-biased gene expression is not the only biological question in my
> experiment. Is the massive size of this effect going to affect the
> "detectability" of other smaller effects? (through normalization or
> correction for multiple testing or other?)
>
> Thanks,
> paolo
>
>
> --
> Paolo Innocenti
> Department of Animal Ecology, EBC
> Uppsala University
> Norbyvägen 18D
> 75236 Uppsala, Sweden
>
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