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  • OneTaq® Hot Start 2X Master Mix with Standard Buffer


    OneTaq® Hot Start 2X Master Mix with Standard Buffer is an optimized blend of Taq and Deep VentR ™ DNA Polymerases combined with an aptamer-based inhibitor. This enzyme blend is ideally suited to routine PCR applications from a variety of templates, including pure DNA solutions, bacterial colonies and cDNA products. The 3´→5´ exonuclease activity of Deep Vent DNA Polymerase increases the fidelity and robust amplification of Taq DNA Polymerase (1). The hot start nature of the enzyme offers convenience with decreased interference from primer dimers and secondary products. The convenient master mix formulation contains dNTPs, MgCl2 and other buffer components and stabilizers listed below, requiring only the addition of primers and DNA template for robust amplification

    Amplification of a selection of sequences with varying GC content from human and C. elegans genomic DNA using OneTaq Hot Start 2X Master Mix with Standard Buffer. The presence of absence of an extended room temperature incubation does not affect performance. Amplicon sizes are indicated next to gel, and GC content is indicated below gel. Marker M is the 1 kb DNA Ladder (NEB #N3232 ).

    Product Source

    An E. coli strain that carries the Taq DNA Polymerase gene from Thermus aquaticus YT-1 and an E. coli strain that carries the Deep VentR DNA Polymerase gene from Pyrococcus species GB-D.

    Advantages and Features


    • High Throughput PCR
    • High Sensitivity PCR
    • Routine PCR
    • AT-rich PCR
    • Colony PCR
    • Long PCR (up to ~6 kb genomic)

    Properties and Usage

    Storage Temperature


    Buffer Composition

    20 mM Tris-HCl
    22 mM KCl
    22 mM NH4Cl
    1.8 mM MgCl2
    5% Glycerol
    0.06% IGEPAL® CA-630
    0.05% Tween® 20
    0.2 mM dNTPs
    25 units/ml OneTaq Hot Start DNA Polymerase
    pH 8.9@25°C

    Heat Inactivation


    Unit Assay Conditions

    1X ThermoPol® Reaction Buffer, 200 μM dNTPs including [3H]-dTTP and 200 μg/ml activated Calf Thymus DNA.

    Quality Control

    Quality Control Assays

    The following Quality Control Tests are performed on each new lot and meet the specifications designated for the product. Individual lot data can be found on the Product Summary Sheet/Datacard or Manual which can be found in the Supporting Documents section of this page.
    • PCR Amplification (DNA Polymerase):

      The polymerase master mix is tested in a polymerase chain reaction (PCR) using a control template and specific primers, resulting in the expected product.


    1. Product specifications for individual components in the OneTaq Hot Start 2X Master Mix with Standard Buffer are available separately.
    2. OneTaq Hot Start 2X Master Mix with Standard Buffer is stable for fifteen freeze-thaw cycles when stored at -20°C.
    3. OneTaq Hot Start 2X Master Mix with Standard Buffer is also stable for one month at 4°C, so for frequent use, an aliquot may be kept at 4°C.


    1. Barnes, W.M (1994). Proc.Natl.Acad.Sci.USA. 91, 2216-2220.
    2. Saiki, R.K. et al. (1994). Science. 91, 2216-2220.
    3. Powell, L.M. et al. (1987). Cell. 50, 831-840.
    4. Sun, Y, Hegamyer, G. and Colburn, N. (1993). Biotechniques. 15, 372-374.

    Supporting Documents

    Material Safety Datasheets

    The following is a list of Material Safety Data Sheets (MSDS) that apply to this product to help you use it safely. The following file naming structure is used to name these document files: [Product Name] MSDS. For international versions please contact us at info@neb.com.


    The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.
    1. How should I set up a PCR using the OneTaq® Hot Start Master Mixes?
    2. Can I use my regular Taq-based cycling conditions for OneTaq® Hot Start DNA Polymerase based products?
    3. How do I activate OneTaq® Hot Start Polymerase?
    4. What are the stability and storage requirements of the OneTaq® Master Mixes?
    5. Can OneTaq® DNA Polymerase be used in colony PCR?
    6. What type of DNA ends result from a primer extension reaction or a PCR using OneTaq® DNA Polymerase?
    7. How long a product can be made by OneTaq® DNA Polymerase?
    8. Can OneTaq® DNA Polymerase be used with uracil-containing primers or bisulfite-treated DNA?
    9. What is the fidelity of OneTaq® DNA Polymerase?
    10. Where can I find help troubleshooting my PCR?
    11. How should I determine an appropriate annealing temperature for my reaction?
    12. How is OneTaq DNA Polymerase different from LongAmp™ Taq DNA Polymerase?
    1. Protocol for OneTaq® Hot Start 2X Master Mix with Standard Buffer (M0484)

    Selection Tools

    Usage Guidelines & Tips

    Troubleshooting Guides

    Interactive Tools

    Application Notes


    • Gorski L, Walker S, Liang AS, Nguyen KM, Govoni J, et al. (2014) Comparison of Subtypes of Listeria monocytogenes Isolates from Naturally Contaminated Watershed Samples with and without a Selective Secondary Enrichment PLoS ONE 9(3), e92467. PubMedID: 24651315, DOI: 10.1371/journal.pone.0092467
    Try using a master mix with OneTaq Hot Start DNA Polymerase to achieve highly specific and sensitive PCR amplification.