Phusion® High-Fidelity DNA Polymerase

High-Fidelity DNA Polymerases are important for applications in which the DNA sequence needs to be correct after amplification. Its unique structure, a novel Pyrococcus-like enzyme fused with a processivity-enhancing domain, increases fidelity and speed. Phusion DNA Polymerase is an ideal choice for cloning and can be used for long or difficult amplicons. With an error rate > 50-fold lower than that of Taq DNA Polymerase and 6-fold lower than that of Pyrococcus furiosus DNA Polymerase (1), Phusion is one of the most accurate thermostable polymerases available. Phusion DNA Polymerase possesses 5´→ 3´ polymerase activity, 3´→ 5´ exonuclease activity and will generate blunt-ended products.

Phusion DNA Polymerase is supplied with standard 5X Phusion HF Buffer, as well as 5X Phusion GC Buffer, which can be used for complex or GC-rich templates. Each of these buffers contains MgCl₂ (1.5 mM at the final [1X] reaction concentration). Reactions can also be optimized using the provided DMSO or MgCl₂ solutions.

Product Source

AnE. colistrain that carries the Phusion DNA Polymerase gene.

Reagents Supplied

The following reagents are supplied with this product:

NEB #

Component Name

Component #

Stored at (°C)

Amount

Concentration

M0530S

-20

Phusion^® High-Fidelity DNA Polymerase	M0530SVIAL	-20	1 x 0.05 ml	2,000 units/ml
MgCl₂ solution	B0510AVIAL	-20	1 x 1.5 ml	50 mM
DMSO	B0515AVIAL	-20	1 x 0.5 ml	100 %
Phusion^® HF Buffer Pack	B0518SVIAL	-20	2 x 1.5 ml	5 X
Phusion^® GC Buffer Pack	B0519SVIAL	-20	1 x 1.5 ml	5 X

M0530L

-20

Phusion^® High-Fidelity DNA Polymerase	M0530LVIAL	-20	1 x 0.25 ml	2,000 units/ml
MgCl₂ solution	B0510AVIAL	-20	2 x 1.5 ml	50 mM
DMSO	B0515AVIAL	-20	1 x 0.5 ml	100 %
Phusion^® HF Buffer Pack	B0518SVIAL	-20	6 x 1.5 ml	5 X
Phusion^® GC Buffer Pack	B0519SVIAL	-20	2 x 1.5 ml	5 X

Features

Robust Reactions – Maximal success with minimal optimization
Extreme Fidelity – >50X greater than Taq.
High Speed – Dramatically reduced extension times (10X faster than Pfu)
High Yield – Increased product yield using minimal amount of enzyme
Versatile – Can be used for routine PCR as well as long or difficult templates

Application Features

PCR
Cloning
Long or Difficult Amplification
High-Throughput PCR

Properties & Usage

Unit Definition

One unit is defined as the amount of enzyme that will incorporate 10 nmol of dNTP into acid insoluble material in 30 minutes at 74°C.

Storage Buffer

20 mM Tris-HCl
100 mM KCl
1 mM DTT
0.1 mM EDTA
200 Âµg/ml BSA
50% Glycerol
1X stabilizers
pH 7.4 @ 25°C

Heat Inactivation

5' - 3' Exonuclease

3' - 5' Exonuclease

Yes

Unit Assay Conditions

25 mM TAPS-HCl (pH 9.3 @ 25°C), 50 mM KCl, 2 mM MgCl², 1 mM b-mercaptoethanol, 200 µM dNTPs including [³H]-dTTP and 15 nM primed M13 DNA.

Companion Products

Deoxynucleotide (dNTP) Solution Set
Deoxynucleotide (dNTP) Solution Mix
Magnesium Chloride (MgCl2) Solution
Phusion® High-Fidelity PCR Kit
Phusion® High-Fidelity PCR Master Mix with HF Buffer
Phusion® High-Fidelity PCR Master Mix with GC Buffer
Deoxynucleotide (dNTP) Solution Mix
Phusion® HF Buffer Pack
Phusion® GC Buffer Pack
Detergent-free Phusion® HF Buffer Pack
Detergent-free Phusion® GC Buffer Pack

References

Frey, B. and Suppman, B. (1995). BioChemica. 2, 34-35.
Chester, N. and Marshak, D.R. (1993). Analytical Biochemistry. 209, 284-290.

Protocols

PCR Protocol Phusion® DNA Polymerase

Usage Guidelines

Selection Charts

Web Tools

FAQs

What are the differences between the various available Phusion® products?
My results are not as expected. Where can I find troubleshooting help?
What ends will my PCR products have?
What does exonuclease activity mean for a DNA polymerase?
Does Phusion® High-Fidelity DNA Polymerase exhibit a strand displacement activity?
What are the properties of this polymerase (fidelity, product ends, max amplicon, modified base incorporation, etc.)?
How should I determine the appropriate annealing temperature for my reaction?
What should the final primer concentration be in my PCR?
My template is GC-rich or supercoiled. How can I optimize my product yield using Phusion® High-Fidelity DNA Polymerase?

Troubleshooting

PCR Troubleshooting Guide

Tech Tips

Phusion has different annealing temperature requirements than most PCR enzymes. Please check out NEB's Tm Calculator to help you determine your optimal Phusion annealing temperature.

Citations

Additional Citations

Wrenbeck E.E., Klesmith J.R., Stapleton J.A., Adeniran A., Tyo K.E., & Whitehead T.A (2016) Plasmid-based one-pot saturation mutagenesis Nat Methods 11, 928-930. DOI: 10.1038/NMETH.4029

Quality Control Assay

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

M0530S_L_v1

Certificate of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety Data Sheets

Legal And Disclaimer

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.Phusion® DNA Polymerase was developed by Finnzymes Oy, now a part of Thermo Fisher Scientific. This product is manufactured by New England Biolabs, Inc. under agreement with, and under the performance specifications of Thermo Fisher Scientific.

Licenses/Patents/Disclaimers:
Notice to purchaser: Limited license.
The purchase price of this product includes a limited, non-transferable license under U.S. and foreign patents owned by BIO-RAD Laboratories, Inc., to use this product. No other license under these patents is conveyed expressly or by implication by purchase of this product.

PHUSION® is a registered trademark and property of Thermo Fisher Scientific.
THERMO SCIENTIFIC® is a registered trademark of Thermo Fisher Scientific.

Phusion® High-Fidelity DNA Polymerase | NEB (2024)