PCR Enzyme

Pfu DNA Polymerase

Ultra pure high fidelity Taq DNA polymerase

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Catalog Number｜Packaging

Mat. No	Ref. No	No. of preps
4992760	GEP101-02	500 U, 2.5 U/μl

Description

Pfu DNA Polymerase is a themostable enzyme with a molecular weight of approximately 90 kDa. It catalyzes the polymerization of nucleotides into duplex DNA in the 5'-3' direction in the presence of magnesium and it also exhibits 3'-5' exonuclease (proofreading) activity, resulting in blunt-ended PCR products without 3'-dA overhanges. Pfu DNA Polymerase is recommended for use in PCR and primer extension reactions that require high-fidelity DNA synthesis.

Pfu DNA Polymerase is expressed from E.coli with cloned Pyrococus Furiosis DNA Polymerase gene and purified and separated by multiple column purification. Because Pfu has 3'-5' exonuclease activity, it can proofread in DNA amplification process, while traditional Taq DNA Polymerase cannot. Although other Taq DNA polymerases such as Vent, Deep Vent, Tli, UITma, etc. have proofreading functions, Pfu has the lowest mismatch rate among all Taq DNA polymerases found so far. Pfu DNA Polymerase has better thermal stability than common Taq DNA polymerase, and it can maintain more than 90% activity at 95°C for 1 hour.

One-tube Pfu PCR Mix (China National High-Tech Product Certification)

■ The Pfu PCR Mix has improved specificity and sensitivity of PCR reaction and can amplify complex templates with high GC content, secondary structure and the like. As low as 2 copies of the target template can be amplified, ensuring more accurate experimental results.

■ The unique Pfu MasterMix formula makes the whole reaction system very stable, and the activity will not be affected by repeated freeze-thaw or long-term storage at 4°C.

■ The stable and efficient pre-prepared PCR mix solution can make the operation fast and simple, greatly reducing labor intensity and sampling error. High-performance PCR enhancer and optimizer are also included in the mix, which reduces the requirements on PCR conditions.

■ This product has both dye-containing and dye-free systems. Dye-containing PCR Mix products can be directly electrophoresed after PCR, without adding sample buffer.
Kit Contents
Applications

High fidelity DNA amplification, such as DNA sequencing, gene expression & cloning, gene site-directed mutagenesis, SNP, end repairing, etc.

It can be used for high fidelity amplification of DNA, such as gene expression cloning, site-directed mutation, single nucleotide polymorphism (SNP) analysis and end repairing.
Storage Condition

-30~-15°C for two years.

Store at -20°C.
Important Notes

● The extension of Pfu DNA Polymerase is about 500-1000 bp/min normally. Because Pfu DNA Polymerase with 3’-5’ exonulease activity may degrade the primers, it should be added later than other components, and immediately go to PCR step.

● As Pfu DNA Polymerase has superior thermostability compared to Taq DNA Polymerase, for high-GC templates, temperature of denaturation can be increased to 98°C, which doesn’t have any influence on Pfu DNA Polymerase’s activity.
Storage Buffer

50 mM Tris-HCl (pH 8.2), 0.1 mM EDTA, 1 mM DTT, Stabilizers, 50% Glycerol.

50 mM Tris-HCl (pH8.2)

0.1 mM EDTA

1 mM DTT

50% Glycerol

Stabilizers
Quality Control

Purity up to standard by SDS-PAGE assay. No exogenous nuclease activity is detected. Single copy gene in human genome could be amplified effectively. No significant activity change when stored at room temperature (15-30°C) for one week.

The purity by SDS-PAGE detection is more than 99%; No activity of exogenous nuclease is detected; Single-copy gene in human genome could be amplified effectively; No significant activity change when stored at room temperature for one week.
Unit Definition

One unit of Pfu DNA Polymerase is defined as the amount that incorporates 10 nmol of dNTPs into acid-insoluble material within 30 min at 74°C with activated salmon sperm DNA as the template/primer.

The activity of 1 unit (U) Pfu DNA Polymerase is defined as the amount of enzyme required to incorporate 10 nmol deoxynucleotides into acid-insoluble substances at 74°C within 30 min using activated salmon sperm DNA as a template/primer.