Product Class: Other

Phusion™ High-Fidelity PCR Master Mix with HF Buffer
cloned at NEB recombinant engineered No hifi pcr 3

Product Introduction

For high speed and high performance PCR. Manufactured and quality-controlled at New England Biolabs, Thermo Scientific™ Phusion™ High-Fidelity DNA Polymerase offers both high fidelity and robust performance.

  • 50X higher fidelity compared to Taq
  • Robust reactions - maximal success with minimal optimization
  • Master mix format with HF buffer ideal for a wide range of AT/GC content
Catalog # Size Concentration
M0531S 100 reactions (50.0 µl) 2 X
M0531L 500 reactions (50.0 µl) 2 X

Product Information

Description

High-Fidelity DNA Polymerases are important for applications in which the DNA sequence needs to be correct after amplification. Phusion High-Fidelity DNA Polymerase offers both high fidelity and robust performance, and thus can be used for all PCR applications. 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 polymerase available. Phusion DNA Polymerase possesses 5´→ 3´ polymerase activity, 3´→ 5´ exonuclease activity and will generate blunt-ended products.

Phusion High-Fidelity PCR Master Mix with HF Buffer is a 2X master mix consisting of Phusion DNA Polymerase, deoxynucleotides and reaction buffer that has been optimized and includes MgCl2. All that is required is the addition of template, primers and water.

Figure 1: Fidelity of DNA polymerases Relative to Taq

Bar graph comparing the fidelity of Phusion™ High-Fidelity DNA Polymerase with Pfu DNA Polymerase and Taq DNA Polymerase 

Fidelity assays were performed using a lacI-based method modified from Frey & Suppman, 1995.


Figure 2: Phusion DNA Polymerase generates robust amplification even with short extension times

Gel representing robust amplification by Phusion™ DNA Polymerase, as compared to High Fidelity Pfu and Modified KOD

A 1.2 kb C. elegans genomic amplicon was analyzed using different polymerases. Reactions were set up in duplicate and visualized by microfluidic LabChip analysis. All reactions were conducted according to manufacturer’s instructions using 30 cycles and extension times of either 30 seconds or 2 minutes as indicated.

Product Source

An E. coli strain that carries the Phusion DNA Polymerase gene.
This product is related to the following categories:
Phusion™ High-Fidelity DNA Polymerases,
Master Mixes,
This product can be used in the following applications:
Long Range PCR,
Fast PCR,
High-Fidelity PCR,
Multiplex PCR,
Specialty PCR,
Routine PCR,
PCR

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  • M0531S     -20    
      Phusion™ High-Fidelity PCR Master Mix with HF Buffer M0531SVIAL -20 2 x 1.25 ml 2 X
      DMSO B0515AVIAL -20 1 x 0.5 ml 100 %
  • M0531L     -20    
      Phusion™ High-Fidelity PCR Master Mix with HF Buffer M0531SVIAL -20 10 x 1.25 ml 2 X
      DMSO B0515AVIAL -20 2 x 0.5 ml 100 %

Properties & Usage

Materials Required but not Supplied

Heat Inactivation

No

Unit Assay Conditions

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

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 per as well as long or difficult templates.

Application Features

  • PCR
  • Cloning
  • Long or Difficult Amplification
  • High-Throughput PCR

Product Notes

  1. Products specifications for individual components in the Phusion High-Fidelity PCR Master Mix with HF Buffer are available separately.

References

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

Protocols, Manuals & Usage

Protocols

  1. PCR Using Phusion® High-Fidelity PCR Master Mix with HF Buffer (NEB #M0531)

Usage & Guidelines

Tools & Resources

Selection Charts

Web Tools

FAQs & Troubleshooting

FAQs

  1. What are the differences between the various available Phusion™ products?
  2. My results are not as expected. Where can I find troubleshooting help?
  3. What ends will my PCR products have?
  4. What does exonuclease activity mean for a DNA polymerase?
  5. Does Phusion™ High-Fidelity DNA Polymerase exhibit a strand displacement activity?
  6. What are the properties of this polymerase (fidelity, product ends, max amplicon, modified base incorporation, etc.)?
  7. How should I determine the appropriate annealing temperature for my reaction?
  8. What should the final primer concentration be in my PCR?

Troubleshooting

Tech Tips

Phusion has different annealing temperature requirements than most PCR enzymes. Please check out www.neb.com/tmcalculator to help you determine your optimal Phusion annealing temperature.

Quality, Safety & Legal

Quality Assurance Statement

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]

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 DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

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.

Licenses

Phusion™ DNA Polymerase was developed by Finnzymes Oy, now a part of Thermo Fisher Scientific. 
PHUSION™ and THERMO SCIENTIFIC™ are trademarks and property of Thermo Fisher Scientific.


Notice to purchaser: 
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 to the purchaser by the purchase of this product.

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