detecting alternative splicing in 3' UTR region, please? - (Apr/13/2007 )
I would like some help please? I am trying to detect alternative transcripts at the 3'UTR region of my gene by RT-PCR. I am wondering if you can design reverse primers at the polyA tail, with forward primers in the penultimate exon and this would detect alternative transcripts. Does anyone know if this can be done? and how to design this type of primer?
Any help & suggestions would be greatly appreciated!
That sounds good but is practically difficult it will take alot of As to make the poly A primer Tm match your 3' coding region primer's TM many race kits overcome this by annealing an adapter to the end of the mRNA and using the adapter primer to PCR between the end of the mRNA and the gene specific primer in the coding region. there are 3' RACE kits that can help with this procedure.
Thank you very much for your help. That's a really good idea that I didn't think of. Fingers crossed it will work!
Best wishes, Lorna