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Method de detects different polyadenylated mRNA isoform in a gene


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#1 cuca

cuca

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Posted 17 June 2009 - 02:13 AM

Hi everybody

My gene has 7 polyadenylation site in the 3'UTR. I would like to know which mRNA PolyA forms are present in my cells and if upon treatment some of them are changed. Does anybody know a technic I could use for that? I am not really in the RNA field and I don't know what I should pay attention but instinctively I had some idea.

Idea 1: PCR located in middle of the tails
Idea 2: PCR using poly dT as revers primer.
Idea 3: 3'RACE, but I don't know what is the advantage compared to normal PCR

Problem 1: the polyA tail can have several lenght
Problem 2: some polyA site are near to each other and there is no ideal primer site in it.
Problem 3: as 3'UTR is composed of untranslated region my primer may recognize a lot of other sequence in the genome.

Do you have maybe a better Idea? What should I pay attention.

Thanks
Cuca




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