T4 DNA Polymerase

#3012

 

100 units

#3012L

 

500 units

DESCRIPTION
T4 DNA Polymerase catalyzes the 5'3' polymerization of DNA from a primed single-stranded DNA template. This enzyme has a highly active 3'5' proofreading exonuclease activity but lacks 5'3' exonuclease activity.

SOURCE
Recombinant strain of E. coli.

APPLICATIONS
Filling-in or labeling the recessed 3' termini created by restriction enzyme digestion of DNA.
3'-end labeling of DNA.
Conversion of termini of double-stranded DNA to blunt-ended molecules.
Labeling DNA fragments for use as hybridization probes.

UNIT DEFINITION
One unit is defined as the amount of enzyme required to catalyze the incorporation of 10nmoles of total nucleotide into an acid insouble form in 30 minutes at 37.

STANDARD UNIT ASSAY CONDITIONS
10mM Tris-HCl (pH7.9), 50mM NaCl, 10mM MgCl2, 1mM DTT, 33M dATP, dCTP and dGTP, 33M [3H-dTTP], 70g/ml denatured calf thymus DNA, and 170g/ml BSA.

CONCENTRATION: 1-3 units/l.

10X REACTION BUFFER
100mM Tris-HCl (pH7.9), 500mM NaCl, 100mM MgCl2, 10mM DTT. Supplement with 50g/ml BSA. dNTPs not included.

STORAGE CONDITIONS
100mM Potassium phosphate (pH6.5), 10mM 2-mercaptoethanol and 50% glycerol. Store at -20.

REFERENCES
1. Kunkel, T. A. et al. (1987) Methods Enzymol. 154, 367.
2. Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, second ed., Cold Spring Harbor Laboratory, Cold Spring Harbor.