单位定义

One unit is defined as the amount of enzyme required to give 50% ligation of HindIII fragments of λ DNA (5´ DNA termini concentration of 0.12 µM, 300- µg/ml) in a total reaction volume of 20 μl in 30 minutes at 16°C in 1X T4 DNA Ligase Reaction Buffer.

Concentration:

400,000 cohesive end units/ml and 2,000,000 cohesive end units/ml

反应条件

1X T4 DNA Ligase Reaction Buffer

Incubate at 16°C

1X T4 DNA Ligase Reaction Buffer

50 mM Tris-HCl

10 mM MgCl2

1 mM ATP

10 mM DTT

(pH 7.5 @ 25°C)

贮存溶液

10 mM Tris-HCl

50 mM KCl

1 mM DTT

0.1 mM EDTA

50% Glycerol

pH 7.4 @ 25°C

热失活

65°C for 10 min