SpeedyLink Ligation Kit enables sticky-end or blunt-end ligation in only 5 minutes at room temperature (25°C). Fast ligation is based on the combination of T4 DNA Ligase and specially formulated buffer. It increases efficiency of ligation to that obtained with T4 DNA Ligase in a standard 1 hour ligation. After 5 min incubation DNA is ready to transform into competent cells.
- Cloning into vectors
- Library construction
- TA cloning
- Linker ligation
- Recircularization of linear DNA
2 x Ligation Buffer:
60 mM Tris-HCl (pH 7.5 at 25°C), 20 mM MgCl2, 20 mM dithiothreitol, 14% PEG 6000, 2 mM ATP.
Avoid multiple cycles of freezing/thawing of the reaction buffer. Thawing should be performed at temperatures not exceeding 10oC. Recommended procedure is to divide the provided reaction buffer into smaller portions. DTT in the Reaction Buffer may precipitate upon freezing, if this occurs vortex the buffer until the precipitate dissolves.
Enzyme Storage Buffer:
10 mM Tris-HCl (pH 7.5 at 22°C), 1 mM DTT, 50 mM KCl and 50% (v/v) glycerol.