TurboLigationTM Kit enables quick ligation of both blunt-end or cohesive-end of DNA fragments in only 5 minutes at room temperature. The kit combines TurboLigaseTM, an ultra-pure recombinant T4 DNA Ligase, and 5X TurboLigation Buffer,
an optimized quick ligation buffer. A 5 minutes reaction is as efficient as a traditional 16℃ overnight ligation reaction. The reaction product is used directly for chemical transformation without any further treatment.
- Adding linkers or adapters to DNA
- Blunt or cohesive end ligation of duplex DNA
- DNA cloning
TurboLigaseTM: 5 unit/µl in Storage Buffer of 10 mM Tris-HCl, pH 7.5, 50 mM KCl, 0.1 mM EDTA, 1 mM dithiothreitol, and 50% glycerol 5X TurboLigation Buffer: 250 mM Tris-HCl, pH 7.5, 50 mM MgCl2, 5 mM dithiothreitol, 5 mM ATP, 25%
polyethylene glycol 8000
4. Stability and Storage
TurboLigase and 5X TurboLigation Buffer are stable at -20℃ for 12 month from date of receipt.
Store TurboLigase and 5X TurboLigation Buffer at -20℃.
Shipped in gel packs.
1. TurboLigaseTM Unit Definition
0.015 Weiss unit (1 cohesive end ligation unit) is defined as the amount of TurboLigaseTM required to give 50% ligation of HindIII fragments of l DNA (5´ DNA termini concentration of 0.15 µM) in a total reaction volume of 20 µl in 30 minutes at 16°C in a buffer of 50 mM Tris-HCl, pH 7.5, 10 mM MgCl2, 1 mM ATP, 10 mM Dithiothreitol.
2. TurboLigaseTM Specific Activity and Purity
TurboLigase has a specific activity >12,000 unit/mg.
TurboLigase is purified to homogeneity.
1. Ligation Reaction
- Warm 5X TurboLigation Buffer to room temperature (22℃-25℃) before setting up TurboLigation reactions.
- Keep TurboLigase at -20℃.
- Thaw competent cells (not included) on ice immediately before setting up TurboLigation reactions.
- Always include a control reaction with vector only.
Set up ligation reaction in 10 µl
5X TurboLigation Buffer 2 µl
vector µl total vector and insert volume up to 7 µl
insert µl use ~1:3 molar ratio of vector to insert
adjust volume with water to 9 µl
TurboLigase 1 µl adding TurboLigase the last increases ligation efficiency
- Mix gently and incubate at room temperature for 5 minutes. Incubation between 5 to 10 min gives the best result.
- Chill the reaction on ice for 1 minute.
- Use 1-3 µl of the ligation reaction for transformation of 20-50 µl competenet cells.