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  • RNase H

    Description

    RNase H (Ribonuclease H ) is an endoribonuclease that specifically hydrolyzes the phosphodiester bonds of RNA which is hybridized to DNA. This enzyme does not digest single or double-stranded DNA.

    Product Source

    An E. coli strain that carries the cloned RNase H gene (rnh) from Escherichia coli.

    Reagents Supplied

    The following reagents are supplied with this product:

    Store at (°C)Concentration
    RNase H Reaction Buffer-2010X

    Properties and Usage

    Unit Definition

    One unit is defined as the amount of enzyme that will hydrolyze 1 nmol of the RNA in [3H]-labeled poly(rA).poly(dT), to acid-soluble ribonucleotides in a total reaction volume of 50 μl in 20 minutes at 37°C in 1X RNase H Reaction Buffer with 10 nmol [3H]-labeled poly(rA) and 12.5 μg poly(dT).

    Reaction Conditions

    1X RNase H Reaction Buffer
    Incubate at 37°C

    1X RNase H Reaction Buffer:
    50 mM Tris-HCl
    75 mM KCl
    3 mM MgCl2
    10 mM DTT
    pH 8.3 @ 25°C

    Storage Temperature

    -20°C

    Storage Conditions

    20 mM Tris-HCl
    100 mM KCl
    10 mM MgCl2
    0.1 mM DTT
    0.1 mM EDTA
    50% Glycerol
    pH 7.5 @ 25°C

    Heat Inactivation

    65°C for 20 min

    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.
    • RNase Activity (1 Hour Digestion):
      The product is tested in a reaction containing a RNA substrate.  After incubation for 1 hour there is no detectable degradation of the RNA substrate as determined by gel electrophoresis.
    • Single-Stranded DNase Activity (Radioactivity Release):
      The product is tested in reaction containing radiolabeled single-stranded DNA. After incubation the exonuclease activity is determined by the % release of radioactive nucleotides.

    References

    1. Davis, R. et al. (1988). Cell Biol.. 8, 4745-4755.
    2. Gubbler, U. and Hoffman, B.J. (1983). Gene. 25, 263-269.
    3. Donnis-Keller, H. (1979). Nucl. Acids Res.. 7, 179.
    4. Goodwin, E.C. and Rottman, F.M. (1992). Nucl. Acids Res.. 20, 916.

    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. Is RNase H active in NEBuffer 1-4?
    2. Is DNA Polymerase I active in RNase H buffer?
    3. Which side of the RNA base does RNase H cut?
    4. What is the activity of RNase H at 30°C?