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Q5 High Fidelity DNA Polymerase

Fidelity at its finest

Q5® High-Fidelity DNA Polymerase sets a new standard for both fidelity and robust performance. With the highest fidelity amplification available (>100 times higher than Taq), Q5 DNA Polymerase results in ultra-low error rates. Q5 DNA Polymerase is composed of a novel polymerase that is fused to the processivity-enhancing Sso7d DNA binding domain, improving speed, fidelity and reliability of performance.

Advantages

  • Highest fidelity amplification (~280x higher than Taq)
  • Ultra-low error rates
  • Superior performance for a broad range of amplicons (from high AT to high GC)
  • Hot start and master mix formats available

The Q5 buffer system is designed to provide superior performance with minimal optimization across a broad range of amplicons, regardless of GC content. For routine or complex amplicons up to ~65% GC content, Q5 Reaction Buffer provides reliable and robust amplification. For amplicons with high GC content (>65% GC), addition of the Q5 High GC Enhancer ensures continued maximum performance. Q5 and Q5 Hot Start DNA Polymerases are available as standalone enzymes, or in a master mix format for added convenience. Master mix formulations include dNTPs, Mg++ and all necessary buffer components.

Comparison of High-Fidelity Polymerases:

1 We continue to investigate improved assays to characterize Q5’s very low error rate to ensure that
we present the most accurate fidelity data possible (Potapov, V. and Ong, J.L. (2017) PLoS
ONE. 12(1): e0169774.).

2 PCR-based mutation screening in lacZ (NEB), lacI (Agilent) or rpsL (Life)
3 Takagi et al (1997) Appl. Env. Microbiol. 63, 4504-4510.

Q5 Hot Start High-Fidelity DNA Polymerase

In contrast to chemically modified or antibody-based hot start polymerases, NEB’s Q5 Hot Start utilizes a unique synthetic aptamer. This molecule binds to the polymerase through non-covalent interactions, blocking activity during the reaction setup. The polymerase is activated during normal cycling conditions, allowing reactions to be set up at room temperature. Q5 Hot Start does not require a separate high temperature activation step, shortening reaction times and increasing ease-of-use. Q5 Hot Start Polymerase is an ideal choice for high specificity amplification and provides robust amplification of a wide variety of amplicons, regardless of GC content.

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Details

Figure 1:  Robust amplification with Q5 (A) and Q5 Hot Start (B) High-Fidelity DNA Polymerases, regardless of GC content

Amplification of a variety of human genomic amplicons from low to high GC content using either Q5 or Q5 Hot Start High-Fidelity DNA Polymerase. Reactions using Q5 Hot Start were set up at room temperature. All reactions were conducted using 30 cycles of amplification and visualized by microfluidic LabChip® analysis.

Figure 2:  Amplification Performance Across a Wide Range of Genomic Targets

PCR was performed with a variety of amplicons, with GC content ranging from high AT to high GC, with Q5 and several other commercially available polymerases. All polymerases were cycled according to manufacturer’s recommendations, including use of GC Buffers and enhancers when recommended. Yield and purity of reaction products were quantitated and represented, as shown in the figure key, by dot color and size. A large dark green dot represents the most successful performance. Q5 provides superior performance across the range of GC content.

Figure 3: Q5 DNA Polymerase offers superior amplification for a wide range of templates, regardless of GC content

Amplification of a variety of human genomic amplicons from low to high GC content demonstrates the broad performance of Q5 High-Fidelity DNA Polymerase. All reactions were conducted using 20 ng of input template and included 30 cycles of amplification. Results were visualized by microfluidic LabChip® analysis. Competitor polymerases were cycled according to manufacturer’s recommendations. For the final three amplicons, GC Buffers or enhancers were used when supplied with the polymerase.

Figure 4: Q5 Hot Start DNA Polymerase offers superior amplification for a wide range of templates, regardless of GC content Amplification

Amplification of a variety of human genomic amplicons from low to high GC content demonstrates the broad performance of Q5 Hot Start High-Fidelity DNA Polymerase. All reactions were set up at room temperature using 20 ng of input template and included 30 amplification cycles. Results were visualized by microfluidic LabChip® analysis. Competitor polymerases were cycled according to manufacturer’s recommendations. For the final three amplicons, GC Buffers or enhancers were used when provided with the polymerase.

Table: Comparison of HighFidelity DNA Polymerases

1 PCR-based mutation screening in lacZ (NEB), lacI (Agilent) or rpsL (Life)
2 Takagi et al (1997) Appl. Env. Microbiol. 63, 4504-4510.
3 Template not specified.
4 Simple templates include plasmid, viral and E. coli genomic DNA. Complex templates include plant, human and other mammalian genomic DNA.
5 Values provided by individual manufacturers.

Available Products

PRODUCT REF. SIZE PRICE
Q5 High Fidelity DNA Polymerase M0491S/L 100/500 units NEB Shop_icon
Q5 High Fidelity 2X Master Mix M0492S/L 100/500 units NEB Shop_icon
Q5 Hot Start High Fidelity DNA Polymerase M0493S/L 100/500 units NEB Shop_icon
Q5 Hot Start High Fidelity 2X Master Mix M0494S/L/X 100/500/500 rxn NEB Shop_icon
Also available as Q5 High-Fidelity PCR Kit:
Q5 High-Fidelity PCR Kit E0555S/L 50/200 rxns NEB Shop_icon
Also avalable as Next-Gen-Seq optimized formula:
NEBNext High Fidelity 2X PCR Master Mix M0541S/L 50/250 rxns NEB Shop_icon

Also available:

PRODUCT REF. SIZE PRICE
Deoxynucleotide Solutions, Mix N0447S 8 µmol each NEB Shop_icon
Deoxynucleotide Solutions, Mix N0447L 40 µmol each NEB Shop_icon

Phusion® is a registered trademark and property of Thermo Fisher Scientific.  Phusion® DNA Polymerase was developed by Finnzymes Oy, now a part of Thermo Fisher Scientific. AccuPrime™ is a trademark of Life Technologies, Inc. Platinum® is a registered trademark of Life Technologies, Inc. PfuUltra™ is a trademark of Agilent Technologies, Inc. Q5®, Deep Vent™ and 9°N™ are trademarks of New England Biolabs, Inc. Vent™ is a registered trademark of New England Biolabs, Inc. LabChip® is a registered trademark of Caliper Life Sciences, part of Perkin Elmer, Inc.

Further information can be found in our Technical Resources section or at neb.com