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  • Deep VentR™ DNA Polymerase

    Description

    Deep VentR DNA Polymerase is the second high-fidelity thermophilic DNA polymerase available from New England Biolabs. The fidelity of Deep VentR DNA Polymerase is derived in part from an integral 3´→ 5´ proofreading exonuclease activity. Deep VentR is even more stable than VentR at temperatures of 95 to 100°C.

    Deep Vent DNA Polymerase crystals (Ira Schildkraut and Rebecca Kucera, New England Biolabs, Inc.)
    Amplification of Jurkat genomic DNA with Deep Vent DNA Polymerase. Amplicon sizes are indicated below gel. Marker M is the 1 kb DNA Ladder (NEB #N3232 ).

    Highlights

    • Isolated from a recombinant source
    • Supplied with 10X Reaction Buffer and 100 mM MgSO4
    • High-Fidelity: 5X greater than Taq
    • Extremely High Thermostability: half-life of 23 hours at 95°C
    • Difficult Templates: ideal for GC-rich or looped sequences

    Product Source

    An E. coli strain that carries the Deep Vent DNA Polymerase gene from Pyrococcus species GB-D. The native organism was isolated from a submarine thermal vent at 2,010 meters (1) and is able to grow at temperatures as high as 104°C.

    Reagents Supplied

    The following reagents are supplied with this product:

    Store at (°C)Concentration
    Magnesium Sulfate (MgSO4) Solution-20100 mM
    ThermoPol® Reaction Buffer-2010X

    Advantages and Features

    Features

    • Ideal for GC-rich or looped sequences

    Applications

    • Primer extension
    • PCR

    Properties and Usage

    Unit Definition

    One unit is defined as the amount of enzyme that will incorporate 10 nmol of dNTP into acid insoluble material in 30 minutes at 75°C.

    Reaction Conditions

    1X ThermoPol® Reaction Buffer

    1X ThermoPol® Reaction Buffer:
    20 mM Tris-HCl
    10 mM (NH4)2SO4
    10 mM KCl
    2 mM MgSO4
    0.1% Triton® X-100
    pH 8.8 @ 25°C

    Storage Temperature

    -20°C

    Storage Conditions

    10 mM Tris-HCl
    100 mM KCl
    1 mM DTT
    0.1 mM EDTA
    50% Glycerol
    0.1% Triton® X-100
    pH 7.4 @ 25°C

    Heat Inactivation

    No

    Molecular Weight

    Theoretical: 89000 daltons

    5' - 3' Exonuclease

    No

    3' - 5' Exonuclease

    Yes

    Strand Displacement

    ++

    Resulting Ends

    • Blunt Ends

    Unit Assay Conditions

    1X ThermoPol Reaction Buffer, 200 µM each dNTP including [3H]-dTTP, 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.
    • Endonuclease Activity (Nicking):
      The product is tested in a reaction containing a supercoiled DNA substrate. After incubation for 4 hours the percent converted to the nicked form is determined by agarose gel electrophoresis.
    • Protein Purity (SDS-PAGE):
      The physical purity is assessed by comparing contaminating protein bands in a concentrated sample to the protein of interest band in a sample of known dilution. The purity is determined by SDS-PAGE.

    Notes

    1. BSA is not provided with this enzyme since its presence is not necessary for most primer extension reactions. However, it is available as NEB #B9001. Acetylated BSA should not be used for primer extension reactions.
    2. The calculated half-life of Deep VentR DNA Polymerase at 95°C is 23 hours.
    3. Diluent D is also available (NEB# B8004). This diluent is recommended for making dilutions of Deep VentR Polymerase.
    4. Additional ThermoPol Reaction Buffer packs for this product are also available (NEB# B9004). Each buffer pack contains 4 vials of 10X buffer (1.5 mls each) and 1 vial of 100 mM MgSO4.

    References

    1. Jannasch, H.W. et al. (1992). Appl. Environ. Microbiol.. 58, 3472-3481.

    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.

    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. Can Deep Vent DNA Polymerase be used in other buffers?
    2. I can't get Deep Vent DNA Polymerase to work yet Taq DNA Polymerase works fine.
    3. Are the DNA fragments produced by Deep Vent DNA Polymerase blunt-ended or do they have the single-base 3' overhang that Taq DNA Polymerase yields?
    4. Can I use Deep Vent DNA Polymerase to polish the ends of DNA fragments?
    5. I want to clone primer extension products made with Vent DNA Polymerase or Deep Vent DNA Polymerase. Are some protocols better than others?
    6. Can Vent DNA Polymerase(NEB# M0254) or Deep Vent DNA Polymerase (NEB# M0258) be used to create blunt ends on primer extension products made with Taq DNA Polymerase (NEB# M0267)?
    1. Guidelines for PCR Optimization for Deep Vent DNA Polymerase
    2. Protocol for a Routine Deep Vent PCR

    Selection Tools

    Usage Guidelines & Tips

    Troubleshooting Guides

    Interactive Tools

    The amount of enzyme in the reaction is critical. Try a final concentration of 0.01 U/µl Deep Vent DNA Polymerase in the reaction.