I have to use NdeI and SacI for double digestion in buffer 4.. Is it feasible??? I m using a peT28a(+) vector?

Question

I have to use NdeI and SacI for double digestion in buffer 4.. Is it feasible??? I m using a peT28a(+) vector?

Answer 1

no....use a Z420b(-) vector....yep.

:D

Answer 2

In the olden days, when we had high, medium, low-salt, and Sac buffers, this would be a simple question.  In 2008, when manufacturers provide different buffers from each other in which different combinations of enzymes work surprisingly well together - I'd have to know which manufacturer's buffer 4 you were talking about, and would have to look it up on their website.  Then I'd have to look at a pET28 map and maybe ask you to clarify whether these were adjacent sites in the multiple cloning site or sites in your insert.  If they were close to each other in the multiple cloning site, I'd probably look up whether they were far enough from each other to get efficient double-cutting.  Depending on how organized your lab is, you may have hard copies of the buffer info for both enzymes sitting around - just read them to see if buffer 4 is good for both. :)