I have been battlin with two digestion reactions and could use some help. I'm trying to cut two pCD vectors that is 6525 base pairs long.
Reaction 1: Xho1/BbvC1
Reaction 2: Xho1/Xba1
I'm using enzymes from NEB. I've tried using both Cutsmart buffer and 2.1 buffer, both of which are compatible with these enzymes. I've tried running the reaction for 1 hour at 37oC and have even gone up to doing an overnight digestion. My cut out are 350 and 480 base pairs, respectively. I've cut up to 2 ug of DNA and still don't see a cut out.
If anyone had a similar problem with these or can provide any expertise in using these enzymes, I would appreciate the insight. Thanks.