Protocol for Cleavage of Mismatched DNA (T:T, G:G, T:G) (NEB #M0678)*
- 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)
to 8 pmol Mismatch Endonuclease I
to 50 μl
- Gently mix the reaction by pipetting up and down and microfuge briefly.
- Incubate at 37° C for 30 minutes.
- 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).