[BioC] GCRMA backgrounds?

Matthew Hannah Hannah at mpimp-golm.mpg.de
Thu Jul 22 09:29:44 CEST 2004

I should have said it was just a logical guess.

What I meant was that if you had 2 homologous genes, obviously it 
is going to be harder to avoid homologous regions if you need to find 
50bp versus 25bp? But this is refering to cross-hybridisation between
PM and related sequences, I don't know how it would affect non-specific 
binding of PM to non-complementary sequences (am I right to distinguish 
these?). I should have said 'less-' rather than non-homologous, or just
dropped the 'non-' in the initial post. Also this would only apply where
there were related sequences present, but then different probe-lengths 
for different sequences wouldn't be ideal.

Also while we're on logic another reason to consider is that with 11-20
probesets per mRNA, for short mRNAs there is already some overlap, this
would be worse for longer probes, making them less independent. It would 
also extend the probed region further from the 3' end from where labelling
occurs and so efficiency may be reduced?

If you need a reference I'm sure the affy website or some of their publications
would have something.

Sorry for any confusion.


-----Original Message-----
From: Michael Barnes [mailto:Michael.Barnes at cchmc.org]
Sent: Mittwoch, 21. Juli 2004 19:49
To: Matthew Hannah; bioconductor at stat.math.ethz.ch
Subject: Re: [BioC] GCRMA backgrounds?

What are references for this?


>>> "Matthew  Hannah" <Hannah at mpimp-golm.mpg.de> 07/21/04 12:45PM >>>

As for the 25mers, the obvious thing to take into account is that
as you increase in length it is more likely that non-homologous
probes will bind as it would be more difficult to find sequences
that are gene specific.


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	I've been using GCRMA and the new speedier version (1.1) 
gives different values than the older slower version (1.0). 

	Looking through the bioconductor mails suggests that 
a few other people identified a similar problem, related to a
background not being subtracted. Hopefully people are on the case, 
but this problem seems to have been around since April. I've been 
plugging GCRMA to my colleagues, who are now starting to use it, 
so I hope the problem can be sorted out.

	On a different note, what technical limitations stop 
Affymetrix going for much longer probes than 25 bases? The work 
of Naef and Magnasco, and Wu and Irizarry, highlight the 
limitations of Affy technology due to cross-hybridisation, when 
there are only 25 bases. Pushing upwards to 50 bases will reduce CH, 
but what other factors then come in? 

	My understanding is that the Affy SNP chips have 25 base 
oligos. What is stopping these chips from also having 
cross-hybridisation issues?	

	Best wishes,

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