Protocol for Cleavage of Mismatched DNA (T:T, G:G, T:G) (NEB #M0678)*

1. Set up the following reaction on ice (Please note that enzyme should always be added last):
   

   COMPONENTS	50 μl RXN	FINAL CONC.
   NEBuffer r2.1 (10X)	5 μl	1X
   Mismatch-containing DNA	to 8 pmol
   Mismatch Endonuclease I	1 μl	80 units
   Nuclease-free Water	to 50 μl

2. Gently mix the reaction by pipetting up and down and microfuge briefly.
   
   
3. Incubate at 37° C for 30 minutes.
   
   
4. Heat inactivate at 70 C° for 5 minutes.

* Other mismatches cleavable by Mismatch Endonuclease I (T:I, G:I and G:U DNA mismatches, T:G and T:U DNA:RNA mismatches and thymine strand of T:C mismatches in DNA) may require the addition of more Mismatch Endonuclease I and/or increased incubation times (up to 18 hours).