Dicer Cleavage - Is there a particular spot? (Jun/21/2006 )
Hi everyone, I need some direction here!
I am wondering what makes Dicer cut where it does? If you were to use long dsRNA, how would Dicer know where to cut it to make your desired RNAi molecule?
Any help would be gladly appreciated.
Check your PM. I hope what I sent you can help
Thank you kindly for your help.
What I can make from it is that Dicer doesn't cut at a particular sequence, but instead is more random - so it makes some effective 19nt siRNA's and some ineffective 19nt siRNA's (or 23nt siRNA's, depeding on where dicer cuts).
What I'm actually trying to do is design a shRNA. There are several programs that can design them, but a typical result I get for an insert that will form a shRNA is along the lines of:
Bam HI cut site - sense - loop - antisense - termination signal - Hind III cut site.
If dicer cuts at every 19 or so bp, then I am correct to say that with a shRNA, you aren't really giving Dicer too many options (as there is only a small dsRNA region before the loop appears) - hence almost always making the siRNA's that you desire?