Skip to content Skip to navigation menu

T4 Polynucleotide Kinase, Cloned

Each T4 Polynucleotide Kinase includes the enzyme and 10X Reaction Buffer without ATP. ATP is available separately.

T4 Polynucleotide Kinase, Cloned

An enzyme for the phosphorylation 5' hydroxy ends of ssDNA and dsDNA, RNA, and nucleoside 3´ monophosphates.
  • Phosphorylate 5' hydroxy ends of ssDNA and dsDNA, RNA, and nucleoside 3 monophosphates
  • Highly active enzyme that readily phosphorylates 5' hydroxyl ends of nucleic acids
  • Flexible: Use newly phosphorylated nucleic acids in a variety of applications such as cloning, next-gen sequencing library prep, and preparation of labelled nucleic acids

Size: 3000 units

Item ID P0503K
TBD
Add to basket to request a quote

Product information

T4 Polynucleotide Kinase (PNK) catalyses the transfer of the γ-phosphate from ATP to the 5´ hydroxyl of ssDNA and dsDNA, RNA, and nucleoside 3´ monophosphates. The enzyme also removes the 3´ phosphate from 3´-phosphoryl polynucleotides, deoxyribonucleoside 3´ monophosphates, and deoxyribonucleoside-3´,5´-diphosphates to form a 3´-hydroxyl group.

Applications

  • Labeling of 5´ termini of DNA and RNA with 32P or 33P for DNA sequencing, blot-hybridisation experiments, or transcript mapping using Mung Bean Nuclease, S1 nuclease, or other nucleases.1,2
  • Phosphorylation of oligonucleotide linkers and other DNA or RNA molecules prior to ligation, or for use in ligation amplification reactions with Ampligase® Thermostable DNA Ligase.
  • Preparation of labelled DNA or RNA molecular weight markers for gel electrophoresis and chromatography.

Figure 1. The absence of DNA exonuclease activity in Epicentre's T4 Polynucleotide Kinase. 40-mer oligonucleotides with (5´-PO4) or without (5´-OH) phosphorylated 5´-termini were incubated with the indicated number of units of T4 PNK in 1X T4 PNK Reaction Buffer for 16 hours at 37°C. Electrophoresis of the incubation mixtures in a 15% polyacrylamide/8 M urea gel demonstrates no degradation of the oligonucleotides.

References

  1. Sambrook, J. et al. (1989) in: Molecular Cloning: A Laboratory Manual (2nd ed.), Cold Spring Harbor Laboratory Press, New York.
  2. Chaconas, G. et al. (1980) Meth. Enzymol. 65, 75.

Unit Definition: One unit of T4 Polynucleotide Kinase converts 1 nmol of 32P from [γ-32P]-ATP into an acid-insoluble form in 30 minutes at 37 °C under standard assay conditions.

Storage Buffer: 50% glycerol containing 50 mM Tris-HCl (pH 7.5), 0.1 M NaCl, 0.1 mM EDTA, 0.1% Triton® X-100, and 1 mM DTT.

T4 PNK 10X Reaction Buffer: 330 mM Tris-acetate (pH 7.5), 660 mM potassium acetate, 100 mM magnesium acetate, and 5 mM DTT. A 10 mM solution of ATP for nonisotopic applications is available separately.

Quality Control: T4 PNK is tested in 5´ phosphorylation of nucleic acids and is free of detectable exo- and endonuclease and RNase activities.

Access support

Need some support with placing an order, setting up an account, or finding the right protocol?

Contact us