Using NEB T4 DNA ligase:
- Mix thoroughly, incubate for 30 minutes at room temperature
- Proceed to transformation. Store excess ligation mixture at -20 °C
Using Roche's Rapid DNA Ligation Kit, stored at -20 °C.
Keep the total mass of DNA below 200 ng.
- Add 10 µL of freshly mixed tube #1 (2x T4 DNA ligase buffer) or 2uL of 10x T4 DNA ligase buffer
- Add 1 µL tube #3 (T4 DNA ligase).
- Mix well, incubate for 15 min. at room temperature.
- Proceed to transformation. Store excess ligation mixture at -20 °C.