T4 DNA Polymerase

Downloads

Certificate of Analysis PDF -Current Lot
MSDS PDF -Current Lot     

Reagents Supplied

10X T4 DNA Polymerase Reaction Buffer

Source

Bacteriophage T4 of Escherichia coli

Description

• Exhibits 5'→3' polymerase and 3'→5' exonuclease activities (1, 2)
• Requires the presence of a single-stranded DNA template and a primer
• Exonuclease, stronger than that found in DNA Polymerase I, is more active on single-stranded DNA than on double-stranded DNA
• Ultrapure recombinant enzyme

Applications

• Adds labeled nucleotides to the recessed 3'-ends of DNA fragments
  Exonuclease activity can be used to remove one or a few nucleotides from 3’-end of double–stranded DNA
• Enzyme suitable for:
  -3’ overhang removal to form blunt ends (3,4)
  -5’ fill-in to form blunt ends (3,4)
  -Probe labeling using replacement synthesis (3,4)
  -Single strand deletion subcloning (5)
  -Second strand synthesis in site-directed mutagenesis (6)

Unit Definition

One unit is the amount of enzyme required to incorporate 10 nmol of total nucleotide into acid insoluble form for 30 minutes at 37°C

Reagents Supplied

10X T4 DNA Polymerase Reaction Buffer

Concentration

5 - 10 Units/µl

Assay Conditions

67 mM potassium phosphate (pH 8.8)
6.7 mM MgCl₂
1 mM dithiothreitol
16.6 mM ammonium sulfate
6.7 µM EDTA
167 µg/ml bovine serum albumin
0.033 mM [α-³²P]dATP

Storage Conditions

Store at -20ºC
Product shipped on dry ice 

References

(1) Goulian, M., Lucas, Z.J. and Kornberg, A. (1968) J. Biol. Chem. 243, 627-638
(2) Lehman, I.R. (1981) Enzymes 14, 51-65
(3) Tabor, S. and Struhl, K. (1989) Current Prot. in Mol. Bio. (Ausubel, F.M., et al., eds) pp. 3.5.10-3.5.12
(4) Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989) Mol. Cloning: A Lab Manual, Second Ed. pp. 5.44-5.47
(5) Dale, R., McClure, B. and Houchins, J. (1985) Plasmid 13, 31-40
(6) Kunkel, T.A., Roberts, J.D. and Zakour, R.A. (1987) Methods Enzymol. 154, 367-382