Science Forums Biology Forum Molecular Biology Forum Physics Chemistry Forum

Science Forums Biology Forum Molecular Biology Forum Physics Chemistry Forum (http://www.molecularstation.com/forum/)
-   Molecular Cloning Forum (http://www.molecularstation.com/forum/molecular-cloning-forum/)
-   -   Restriction, Klenow Blunting, and Re-ligation all in one Tube, no Gel Extraction? (http://www.molecularstation.com/forum/molecular-cloning-forum/87723-restriction-klenow-blunting-re-ligation-all-one-tube-no-gel-extraction.html)

carmeyeii 03-15-2013 07:09 PM

Restriction, Klenow Blunting, and Re-ligation all in one Tube, no Gel Extraction?
 
Hi!

Iím trying to make a single HindIII site out of two sites: HindIII and XhoI in a plasmid, for which I need to cut with those two restriction enzymes, blunt the ends with Klenow and religate the plasmid so Iím eft with only the HindIII site.

After the digestion, can I add Klenow, and then ligase all in one tube, i.e., without wasting time on an extra gel extraction step?

Iíve read that Klenow can be added directly to the restiction digestion, but Iím curious as to if one can further re-ligate intramolecularly directly as well.

Thanks for your input!
Carmen


All times are GMT. The time now is 10:11 AM.

Powered by vBulletin® Version 3.8.4
Copyright ©2000 - 2014, Jelsoft Enterprises Ltd.
Copyright 2005 - 2012 Molecular Station | All Rights Reserved

Page generated in 0.08825 seconds with 11 queries