Protocol for Lambda Exonuclease (M0262)
Lambda Exonuclease efficiently degrades 5' phosphorylated linear dsDNA from 5' to 3' direction, leaving supercoiled dsDNA intact.*
1. Set-up the reaction as follows:
|Components||50 μl REACTION
|DNA|| up to 5 μg
| Lambda Exonuclease Reaction Buffer (10X)
|| 5 μl (1X)
| Lambda Exonuclease
|| 1 μl (5 units)
| Nuclease-free H2O
|| up to 50 μl
2. Incubate at 37°C for 30 minutes.
3. Stop reaction by adding EDTA to 10 mM.
4. Heat Inactivation 75°C for 10 minutes.
5. To clean up treated samples, we recommend using one of the following steps:
a. Column clean up (we recommend the Monarch® PCR & DNA Cleanup Kit, NEB #T1030) or
b. Running the reaction on an agarose gel, and then extracting the DNA (we recommend the Monarch Gel Extraction Kit, NEB #T1020), or
c. Performing a phenol/chloroform extraction followed by ethanol precipitation.
*Note: For more precise results we recommend titration of the enzyme to the intended substrate.