Specific targeting of alternative splice forms?
Posted 11 March 2010 - 09:50 AM
given that alternative splice forms all start out on the same piece of pre-mRNA, is it possible to target a particular one? Having trawled through the archives it seems that some people are saying yes, but at the same time it seems that the degradation happens intra-nuclearly, so surely at least some of the prespliced mRNA must be degraded? Can't seem to get the answer from the literature so if anyone does have a reference explaining why it does or doesn't that would be great. I need to know that I'm having zero effect on the splice form I wish to preserve.
It may or may not be important, but I'm planning to transfect in shRNAs to specifically target an exon about 2500bp into a final mRNA of 3kb
Posted 12 March 2010 - 07:44 AM
(1) Morcos PA. Achieving targeted and quantifiable alteration of mRNA splicing with Morpholino oligos. Biochem Biophys Res Commun. 2007 Jun 29;358(2):521-7. Epub 2007 May 7.
(2) Draper BW, Morcos PA, Kimmel CB. Inhibition of zebrafish fgf8 pre-mRNA splicing with morpholino oligos: A quantifiable method for gene knockdown. Genesis. 2001 Jul;30(3):154-6.
(3) Moulton JD, Yan YL. Using Morpholinos to control gene expression. Curr Protoc Mol Biol. 2008 Jul;Chapter 26:Unit26.8.
Edited by Jon Moulton, 12 March 2010 - 07:47 AM.
Gene Tools, LLC
Posted 15 March 2010 - 04:42 AM
Thanks a lot, I'll check these ones out.
Also, I've been using siRNA scales to design my targets. How do these predictions work out in general? I like the fact it gives an estimated level of knock-down, but being that I have had a couple of suggestions of sequences that knock it down by 102%, I have a feeling that they might be taking the predicitive nature of the algorithm a bit too far!