Strange problem with digestion and dephosporylation of PCR product
Posted 19 January 2011 - 09:00 AM
To get 2 PCR products I used two sets of primers with BamHI and Not1 sites and I added extra 6 nucleotides to each primer. After PCR I got my two products exactly the size I wanted (+-1000bp) and no other products. When I digested them with BamH1 and Not1 (1hr, 37 celsius degrees, buffer suitable for both), dephosphorylated them (CIP phosphatase) and purified on gel I got two bands for both products- one the same size as PCR products and one about 50-100bg bigger... Does anyone have any idea? I purified both products and I tried to ligate them twice but my ligations didn't work (I didn't get any colonies).
Posted 20 January 2011 - 02:23 AM
Posted 22 January 2011 - 08:41 PM