[BioC] presence/absence call using oligo array?

[Amy Mikhail]

Hi Xinxia,

Just curious about your message.  It is quite common to look at parasite
genes being expressed at different stages of the parasite lifecycle,
including when the parasite is in the host, as is the case for your
infected tissue.  However I'm a bit confused about the RNA you
cohybridised with, as if there are no parasite RNAs in it you would be
comparing parasite gene expression in your infected tissue with a blank
(did I read correctly that your microarrays have only parasite genes on
them, not host genes?).

Are you using 2-colour arrays?  If so expression levels for each gene are
normally read from one condition relative to the other.  This means that
whatever you co-hybridised with has to have the same set of genes being
expressed at some level - otherwise you have nothing to compare your
condition of interest with.  The results you get in this way are relative
expression levels for each gene of interest, not absolute expression
levels. As I understand it you cannot use microarrays for absolute
expression levels because there are too many factors that can disrupt mRNA
copy number during probe preparation - if you are interested in actual
copy number you would have to do something like quantitative RT-PCR.

Hope this helps,

Regards,
Amy

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Dear Bioc,

Has someone ever used long oligo arrays to test if genes were expressed or

not? I have two microarray slides on which oligonucleotides (65mer) for genes
from a parasite genome were spotted. One gene has one oligo but spotted twice
on each array. One slide has a RNA sample extracted from infected tissues 
(therefore mixture of both parasite RNAs and RNAs from host tissues) 
co-hybridized with RNAs from uninfected host tissue (no parasite RNAs). 
Another is a biological replicate but with dye-swap. People want to know
what
parasite genes are expressed in the infected tissue. Is it possible to do so?
Related publications are highly appreciated.

Best,
Xinxia
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Amy Mikhail
Research student
University of Aberdeen
Zoology Building
Tillydrone Avenue
Aberdeen AB24 2TZ
Scotland
Email: a.mikhail at abdn.ac.uk
Phone: 00-44-1224-272880 (lab)