T3 DNA Ligase

Description:

Pfu DNA Ligase is a heat-stable enzyme that facilitates the formation of phosphodiester bonds between the 5´-phosphate and 3´-hydroxyl groups of two neighboring DNA strands that are fully paired and aligned with a complementary DNA strand, ensuring the absence of gaps. This ligase requires ATP as a cofactor and exhibits activity at elevated temperatures, ranging from 45°C to 70°C.

Pfu DNA Ligase is highly effective in ligating nicks in DNA during incubations at high temperatures. Its extreme thermostability allows it to withstand the demanding conditions of PCR processes. While it does not ligate short 4-base overlaps commonly found in restriction enzyme digests, it is highly efficient at ligating 12-base pair overlaps. This enzyme is isolated from a recombinant source, ensuring its purity and effectiveness for precise molecular biology applications.

Included in the package is a 10X Pfu DNA Ligase Reaction Buffer (10 mM Tris-HCl; 600 µM ATP; 2.5 mM DTT; 2.5 mM MgCl2; 0.1% Triton® X-100).
The enzyme is supplied at a concentration of 40,000 units/ml. (Unit Definition: (Cohesive End Unit) One unit is defined as the amount of enzyme required to give 50% ligation of the 12-base pair cohesive ends of 1 µg of BstEII-digested λ DNA in a total reaction volume of 50 µl for 15 minutes at 45°C. A cohesive end unit is equivalent to the nick-closing unit.)

Applications:

• Pfu DNA Ligase will ligate this substrate:

• Detection of allele-specific genes using Ligase Detection Reaction (LDR) and Ligase Chain Reaction (LCR).
• Incorporation of a phosphorylated oligonucleotide during PCR amplification for mutagenesis.

Source:
Purified from an E. coli strain containing the cloned ligase gene.

Reaction Conditions:
1X Pfu DNA Ligase Reaction Buffer
Incubate at 45°C

Heat Inactivation:
No

Storage:
-20°C

Pack Sizes:
5,000 units (cat. no. PDL-100)
25,000 units (cat. no. PDL-200)