[BioC] sanity check: recipe for p-value from RMA expression
estimates
Reiner Schulz
rschulz at cs.umd.edu
Thu Feb 17 10:30:34 CET 2005
i'd like to repeat what was done as part of:
'Summaries of Affymetrix GeneChip probe level data' by Irizarry et al.
Nucleic Acid Res., 2003, 31(4):e15
http://nar.oupjournals.org/cgi/content/full/31/4/e15
that is, compare the Affymetrix I/D/NC calls/p-values for a differential
expression experiment involving just 2 arrays with those based on the
RMA method.
in the above paper, it says: 'Test statistics can be created for RMA
[...] based on estimates of standard error obtained from probe level
data.' at this point, the authors reference:
'Exploration, normalization, and summaries of high density
oligonucleotide array probe level data' by Irizarry et al. Biostatistics
4:249-264 (2003)
http://biostatistics.oupjournals.org/cgi/content/abstract/4/2/249
it spells out the linear model that is least-squares-fitted to the
measured intensities for the probes in a probe set:
Y = mu + alpha + epsilon, where mu is the expression estimate (log-scale
expression level for the respective array), alpha a probe affinity
effect, and epsilon an independent identically distributed error with
mean 0.
the rma function of the affy R package provides just mu for a probe set,
but not epsilon. function fitPLM of the affyPLM package returns what is
called 'chip level parameter estimates' and 'standard errors'.
i reckon that the latter correspond to mu and epsilon, respectively.
please correct me if i'm wrong here.
given mu and its standard error (SE) for the same probe set, but on 2
different arrays, i'd naively use t = |mu1 - mu2| / (SE1 + SE2) as the
test statistic where df = 2 * number of probes in the probe set - 2.
that provides me w/ a p-value that i can compare to Affymetrix'.
does this make sense, and is this what was done in the paper at the top?
cheers,
Reiner
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