Restriction Digest

Protocol 1. Check the NEB (or other supplier) website to confirm enzyme compatibility 2. In a PCR tube add the reaction components according to the table listed below 3. Incubate at 37°C for 30 min if time-saver qualified, 60 minutes if not

Example Reaction

Component Volume ddH2O Up to 50 µL 10X rCutSmart Buffer 5 µL DNA x (1 µg) Enzyme 1 1 µL Enzyme 2 1 µL

Notes - NEB High-Fidelity (HF) enzymes are ordered whenever possible, they all use rCutSmart reaction buffer and are usually time-saver qualified Hi - NEB has the option to select your specific enzymes and generate a detailed protocol - Heat inactivation may be recommended if there are additional steps in the workflow without a DNA purification step

References, related resources, and acknowledgments - https://nebcloner.neb.com/#!/redigest