T4 DNA Ligase

#3011

#3011HC

 

300 units

#3011L

#3011HCL

 

1,500 units

DESCRIPTION
T4 DNA Ligase catalyzes the formation of a phosphodiester linkage between 5'-phosphoryl group and adjacent 3'-hydroxyl group of duplex DNA or RNA in a blunt-ended or cohesive-ended configuration. ATP is required for the reaction.

SOURCE
Recombinant strain of E. coli.

APPLICATIONS
Joining of double-stranded DNA fragments with cohesive ends or blunt ends.

UNIT DEFINITION
One Weiss unit is defined as the amount of enzyme required to catalyze the exchange of 1nmole of 32P from pyrophosphate to ATP, into Norit-adsorbable material in 20 minutes at 37. The reaction conditions for Weiss unit definition assay are : 66mM Tris-HCl (pH 7.6), 6mM MgCl2, 1mM DTT, 0.066mM ATP and 3.3M [32PPi]. One Weiss unit is equivalent to 67 cohesive end ligation unit which is the amount of enzyme required to give 50% ligation of Lambda DNA Hind III fragments in 30 minutes at 16 in 20l reaction mixture and a 5' DNA termini concentration of 0.12M (300g/ml).

STANDARD UNIT ASSAY CONDITIONS
50mM Tris-HCl (pH7.8), 10mM MgCl2, 10mM DTT, 1mM ATP.

CONCENTRATION: 2-4 units/l, 10-20 units/l (High concentration product)

10X REACTION BUFFER
500mM Tris-HCl (pH7.8), 100mM MgCl2, 100mM DTT, 10mM ATP.

STORAGE CONDITIONS
10mM Tris-HCl (pH 7.4), 50mM KCl, 0.1mM EDTA, 1mM DTT, 0.2mg/ml BSA and 50% glycerol. Store at -20.

REFERENCES
1. Helfman, D.M. et al. (1987) Methods Enzymol. 152, 349. 2. Weiss, B. et al. (1968) J. Biol. Chem. 243, 4543.