Ligation Reaction
Buffers
T4 DNA Ligase Reaction Buffer
Protocol
- Resuspend DNA fragment(s) in 10 μl of TE Buffer.
- Add 10 μl of kinase reaction.
- Add 1.5 μl of 10X ligase buffer (500 mM Tris-HCl (pH 7.5), 100 mM MgCl2, 100 mM DTT, 10 mM ATP).
- Adjust to 23 μl with water.
- Add T4 DNA Ligase 800 units (NEB).
- Incubate at 16°C overnight.
- Heat inactivate reaction 75°C for 5 minutes.