T4 DNA Polymerase

Source

Bacteriophage T4 of Escherichia coli

Description

• T4 DNA Polymerase 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
• T4 DNA Polymerase 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)

Reagents Supplied

10X T4 DNA Polymerase Reaction Buffer

Unit Definition

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

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