pfu DNA polymerase

Product Information

Product description: pfu DNA Polymerase is a recombinant thermostable enzyme from Pyrococcus furiosus, expressed in Escherichia coli. It catalyzes the polymerization of nucleotides into duplex DNA in the 5'→3' direction and exhibits 3'→5' exonuclease activity in the presence of Mg²⁺, can correct the wrong DNA amplification base. The error rates of pfu DNA Polymerase is the lowest, is just 1.3x10^-6. pfu DNA Polymerase is recommended for use in PCR and primer extension reactions that require high fidelity.

Source: Recombinant E.coli

Concentration: 3U/μL

Unit Definition: One unit is defined as the amount of enzyme required to catalyze the incorporation of 10 nmoles of dNTPs into acid insoluble material in 30 minutes at 75℃.

Storage Buffer: 50mM Tris-HCl (pH8.2), 0.1mM EDTA, 1mM DTT, 0.1%(v/v) Tween-20, 0.1%(v/v) NP-40, 50%(v/v) glycerol.

Companion Product: 10X pfu Reaction Buffer, ON-068, 200mM Tris-HCl (pH8.8 at 25℃), 100mM KCl, 100mM (NH₄)₂SO₄, 20mM MgSO₄, 1% Triton X-100.

Key points of operation

1. pfu DNA Polymerase has 3'→5' exonuclease activity, so the extension rate (0.5-0.6kb/minute) is lower than Taq DNA Polymerase(1kb/minute), the extension time should be based on the amplified product length; at the same time, because of the 3'→5' exonuclease activity , pfu DNA Polymerase may degrade the primer, so to add pfu DNA Polymerase to the reaction system at last.

2. Require a higher purity of primers while amplifying with pfu DNA Polymerase. The primer length should be greater than 18 bp, Tm is at 55-80℃, primer concentration is between 0.1-0.5μM, slightly higher than the Taq DNA Polymerase.

3. For high GC content of template, denaturation temperature can be raised to 98 ℃, has no effect on the activity of DNA Polymerase.

Security Information

Storage Conditions: -20℃

Quality Assurance: Free of endonuclease, exonuclease and Nickase activities.

Physical Purity: ＞95% by SDS-PAGE.

Reference

1. Kelly S. Lundberg, Dan D. Shoemaker, Michael W. W. Adams, et al. Gene 108, 1-6 (1991).