Q5® High-Fidelity DNA Polymerases

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  • Fidelity at its Finest

    See what our customers are saying about Q5® High-Fidelity DNA Polymerases

    Q5® High-Fidelity DNA Polymerase (NEB #M0491) 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.

    • Why Choose Q5® High-fidelity Polymerase?

      Not sure why Q5® is your best choice for high-fidelity amplification of GC-rich targets? NEB's scientists will show you why we call Q5 an "ultra-high fidelity polymerase".

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    • Important Tips for Q5® Polymerase

      Here are some quick tips for getting the most out of NEB's Q5® High-Fidelity DNA Polymerase.

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    • How to Amplify GC-rich DNA

      Looking for tips on dealing with GC-bias in DNA amplification? NEB scientists have the expertise you need!

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    • Choose the Right DNA Polymerase for PCR

      Make sure you're using the optimal polymerase for your DNA amplifications. Get tips on choosing the right DNA Polymerase for your application.

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    Comparison of High-Fidelity Polymerases

    1 PCR-based mutation screening in lacZ (NEB), lacI (Agilent) or rpsL (Life)
    2 Due to the very low frequency of misincorporation events being measured, the error rate of high-fidelity enzymes like Q5 is difficult to measure in a statistically significant manner. Although measurements from assays done side-by-side with Taq yield a Q5 fidelity values of approximately 200X Taq, we report “>100X Taq” as a conservative value.
    3 Takagi et al (1997) Appl. Env. Microbiol. 63, 4504-4510.

    Advantages

    • Highest fidelity amplification (>100X 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 (NEB #B9027) 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.

    Robust Amplification with Q5 (A) and Q5 Hot Start (B) High-Fidelity DNA Polymerases

    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.

    In contrast to chemically modified or antibody-based hot start polymerases, NEB's Q5 Hot Start (NEB #M0493) 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.

    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.

    Master Mix and Stand-Alone Formats Provide Convenience and Flexibility

    Q5® is a registered trademark of New England Biolabs, Inc.
    LabChip® is a registered trademark of Caliper Life Sciences, part of Perkin Elmer, Inc.

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    PCR Selection Tool

    Choose from one of the largest selections of polymerases for PCR applications from the leader in enzyme technology and bring unparalleled confidence to your experiments.

    Comparison of High-Fidelity Polymerases

    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.

    Five Quality Features of Q5

    1. Fidelity – the highest fidelity amplification available (>100X higher than Taq)
    2. Robustness – high specificity and yield with minimal optimization
    3. Coverage – superior performance for a broad range of amplicons (from high AT to high GC)
    4. Speed – short extension times
    5. Amplicon length – robust amplifications up to 20 kb for simple templates, and 10 kb for complex

    DNA Polymerase Selection Chart

    NEB offers a guidelines for choosing the correct DNA polymerase for your application by providing a list of specific properites.
    Several factors govern which polymerase should be used in a given application, including: 

    Template/product specificity: Is RNA or DNA involved? Is the 3´ terminus at a gap, nick or at the end of the template? 

    Removal of existing nucleotides: Will the nucleotide(s) be removed from the existing polynucleotide chain as part of the protocol? If so, will they be removed from the 5´ or the 3´ end? 

    Thermal stability: Does the polymerase need to survive incubation at high temperature or is heat inactivation desirable? 

    Fidelity: Will subsequent sequence analysis or expression depend on the fidelity of the synthesized products?