Thermostable Taq DNA Ligase catalyzes the formation of a phosphodiester bond in duplex DNA containing adjacent 5′-phosphoryl and 3′-hydroxyl termini, using NAD+ as a cofactor.
- Ligation of DNA fragments at elevated temperatures (45°C-65°C).
- Ligase Chain Reaction.
- Gibson Assembly of DNA fragments (Gene Synthsis, joning multiple large DNA fragments).
- Taq DNA Ligase, 40 U/µl in storage buffer: 10 mM Tris-HCl, 50 mM KCl, 1 mM Dithiothreitol, 0.1 mM EDTA, 0.1% Tween-20 50% Glycerol, pH 7.5 @ 25°C.
- 10X Taq DNA Ligation Buffer: 200 mM Tris-HCI, 250 mM KCl, 100 mM MgCl2, 5 mM NAD, 0.1% Triton X-100, pH 7.6 @ 25°C
1 unit is defined as the amount of Taq DNA Ligase required to join 50% of 1 µg of the 12-base cohesive ends of Lambda DNA cut with Hind III in 50 µl 1X Taq DNA Ligase Buffer following a 10 minute incubation at 45°C.
- The absence of endo-, exodeoxyribonucleases, phosphatases, and ribonucleases confirmed by appropriate quality tests.
- Functionally tested for the capacity to join cohesive- and blunt-end DNA fragments.