I need help on Ligations

William Bonner wmbonner at helix.nih.gov
Fri Apr 14 20:57:38 EST 1995


akravetz at MO.NET (Andy Kravetz) wrote:

>Jennifer Lyon (movatech at mailbag.com) wrote:
>: NcoI is a tricky cutter on the ends of molecules. If you put an NcoI site on the end ofa PCR product,\
>: make sure you left enough space for NcoI to sit down and cut. I'd recommend giving it about 10 bps. 
>: For almost all retrictin enzymes, its a good idea to leave them about 6 bp's on the end.
>: A good source for cutting info on R.E.'s is NEB's catalog.

>yeah, Jennifer, but I checked the B/M catalog and it said that I did not 
>need to have extra bases. As such, I designed my primer without an extra 
>base. live and Learn right??? So to all, what do I do now.. If truly, the 
>NcoI is not cutting then I am SOL. Any hints...

Blunt clone into a vector like pCR-script (Stratagene) or other, then
cut out with NcoI.-Bill, wmbonner at helix.nih.gov





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