Gene ratio after RT-PCR on a multigene template

mblov2439 at my-deja.com mblov2439 at my-deja.com
Sun Jun 25 09:42:21 EST 2000


I got genes that encodes MHC cl. II beta, in fish MHC cl. II beta is
encoded by 2 - 3 loci(?). To describe the ratio between the loci in
mRNA, I have isolated mRNA, made cDNA by olig-T primer, and done MHC cl.
II beta spesifikk RT-PCR on the cDNA. RT-PCR product is cloned into
plasmids and 20 clones are sequenced. The PCR primers amplify them
all, and there are only a few single nucleotide polymorphisms that
separates the expressed genes. An awfull design with much work for
little results, but what do the ratio of the cloned plasmids say me. The
most expressed mRNA will hopefulle also have the largest ratio of the
plasmids, but will the ratio change during a PCR, and will small
differences become larger or smaller during the PCR.

 What happens in the tube during the PCR, will the ratio between.
What kind of aspects would you like me to focus on, to resolve the
expression pattern.

We have got a quantitative PCR, but I would like to know a little bit of
what happening in the tube during the PCR.

Best regards : Olav V


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