DNA Digest

From 2010.igem.org

50 ul digest

1. Quickly vortex all ingredients (Buffer, BSA, DNA, Enzymes) before beginning.

2. Add the following in a micro-centrifuge tube:

         a.5 ul of Buffer 2
         b.1 ul of BSA
         c.0.5 picomoles of DNA
         d.Water to make 50 ul

3. Vortex Enzymes and add 20 units (1 ul) of each to the tube

4. Incubate reaction in a 37 degrees Celsius water bath for one hour

5. Heat kill the digest for 20 minutes at 80 degrees Celsius.

6. If digesting vector add 1 ul phosphotase and 5 ul of Phosphotase Buffer and incubate an additional 45 minutes.

7. Heat kill enzymes at 80 degrees for 20 minutes.

8. Store digested DNA in the freezer (-20 degrees Celsius).