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  • Hot Start Taq 2X Master Mix

    Description

    Hot Start Taq 2X Master Mix is an optimized ready-to-use solution containing Hot Start Taq DNA Polymerase, dNTPs, MgCl2, KCl and stabilizers. It is ideally suited to routine PCR applications from templates including pure DNA solutions, bacterial colonies, and cDNA products. It is recommended for amplification up to 5 kb.

    Hot Start Taq DNA Polymerase is a mixture of Taq DNA Polymerase and an aptamer-based inhibitor. The inhibitor binds reversibly to the enzyme, inhibiting polymerase activity at temperatures below 45°C, but releases the enzyme during normal cycling conditions allowing reactions to be set up at room temperature. The aptamer-based hot start does not require a separate high temperature incubation step to activate the enzyme. Hot Start Taq DNA Polymerase possesses a 5´→ 3´ polymerase activity(1)(2)(3) and a 5´ flap endonuclease activity(4)(5).

    Product Source

    An E. coli strain that carries the Taq DNA Polymerase gene from Thermus aquaticus YT-1

    Advantages and Features

    Applications

    • High-specificity PCR
    • Routine PCR
    • High-throughput PCR
    • Microarray Analysis
    • Colony PCR

    Properties and Usage

    Storage Temperature

    -20°C

    Buffer Composition

    10 mM Tris-HCl
    50 mM KCl
    1.5 mM MgCl2
    0.2 mM dNTPs
    5% Glycerol
    25 units/ml Hot Start Taq DNA polymerase
    pH 8.6@25°C

    Heat Inactivation

    No

    Unit Assay Conditions

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

    Notes

    1. Hot Start Taq 2X Master Mix is stable for fifteen freeze-thaw cycles when stored at -20°C.
    2. Hot Start Taq 2X Master Mix is also stable for three months at 4°C, so for frequent use, an aliquot may be kept at 4°C.

    References

    1. Chien, A., Edgar, D.B. and Trela, J.M. (1976). J. Bact.. 127, PubMedID: 1550-1557
    2. Kaledin, A.S., Sliusarenko, A.G. and Gorodetskii, S.I. (1980). Biokhimiya. 45, 644-651.
    3. Lawyer, F.C. et al. (1993). PCR Methods and Appl.. 2, 275-287.
    4. Longley, M.J., Bennett, S.E. and Mosbaugh D.W. (1990). Nucleic Acids Res.. 18, 7317-7322.
    5. Lyamichev, V., Brow, M.A. and Dahlberg, J.E. (1993). Science. 260, 778-783.
    6. Sun, Y., Hegamyer, G. and Colburn, N. (1993). Biotechniques. 15, PubMedID: 372-374
    7. Sarkar, G., Kapelner, S. and Sommer, S.S. (1990). Nucleic Acids Res.. 18, 7465.

    Supporting Documents

    Datacards

    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. Does Hot Start Taq 2X Master Mix require an extended initial incubation to activate the polymerase?
    2. How should I set up a PCR using the Hot Start Taq 2X Master Mix?
    3. What type of DNA ends result from a primer extension reaction or a PCR reaction using Hot Start Taq DNA Polymerase?
    4. When should Hot Start Taq DNA Polymerase be used in a primer extension reaction or for PCR?
    5. Will the 5'→3' flap endonuclease activity of Hot Start Taq DNA Polymerase degrade primers?
    6. Can Hot Start Taq DNA Polymerase be used for nick translation?
    7. How should I determine an appropriate annealing temperature for my reaction?
    8. What is the longest amplicon that can be obtained with Hot Start Taq DNA Polymerase?
    9. Can Hot Start Taq DNA Polymerase be used with uracil-containing primers or bisulfite-treated DNA?
    1. Protocol for a PCR reaction using Hot Start Taq 2X Master Mix (M0496)

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