Question: Scaling Before Anova?
gravatar for Rossella
9.4 years ago by
Rossella370 wrote:

Hi, I want to perform an anova analysis for identifying differentially expressed genes. Should I scale the data using quantile normalization or median absolute deviation before looking for differentially expressed genes or should I apply anova directly on data obtained through RMA? I am working with Affymetrix single channel microarrays.

Thanks a lot in advance

data • 3.4k views
ADD COMMENTlink modified 9.4 years ago by Daniel Swan13k • written 9.4 years ago by Rossella370
gravatar for Daniel Swan
9.4 years ago by
Daniel Swan13k
Aberdeen, UK
Daniel Swan13k wrote:

For completeness here's the answer I already posted to your query on StackOverflow

You can apply ANOVA after RMA, since it includes background adjustment, summarisation and quantile normalisation steps (see here). For non-Affy data, something along the lines of log transformation/quantile normalisation or VST (variance stablilising transformation)/RSN (robust spline normalisation) for Illumina data could be used prior to ANOVA.

By the way you will also want to do some filtering on the data after RMA and prior to ANOVA to remove probesets of low variance. If you're working in R you will want to look at genefilter.

ADD COMMENTlink written 9.4 years ago by Daniel Swan13k
gravatar for Michael Dondrup
9.4 years ago by
Bergen, Norway
Michael Dondrup47k wrote:

The RMA preprocessing step most likely already involved normalization, at least if performed by running rma via expresso or justRMA from the affy package.

ADD COMMENTlink written 9.4 years ago by Michael Dondrup47k

Absolutely, it's quantile normalisation, to be precise.

ADD REPLYlink written 9.4 years ago by Laurent1.7k
gravatar for Chris Evelo
9.4 years ago by
Chris Evelo10k
Maastricht, The Netherlands
Chris Evelo10k wrote:

RMA should be enough for normalisation. But you should do some quality control and possibly filtering of bad data before you continue. There are many Bioconductor packages that help you do that We try to offer some guidance on

ADD COMMENTlink written 9.4 years ago by Chris Evelo10k
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