Have been trying for a while now to clone a PCR product into pcDNA3.1 vector.
Am using BamHI and NotI.
Using each enzyme separately I see a cut product when run in a gel along side uncut vector (2.5hr digestions).
However in doing a double digest I only get one cut.
Have tried sequential digest too, using Not I, purifying, then using BamHI. Seems that BamHI no longer wants to cut the linearized vector.
NEB double digest tool says that these enzymes are compatible in NEB3.
Any explanations? Or am I just doing something horribly wrong?
Any advice welcome!!
Edited by Ryang, 07 October 2009 - 01:48 PM.