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  • Nt.AlwI

    This enzyme has transitioned to an improved new buffer system. Visit NEBCutSmart.com for further details.
     
    The new and current Double Digest Finder and current Activity/Performance Chart for the CutSmart buffer system are available. The previous version of the Double Digest Finder, as well as the previous Version of Activity/Performance Chart that use the former buffer system, are still available for your convenience.
    cloned at neb recombinant incubation temp heat inactivation dam
    R0627_TN
    Catalog #SizeConcentrationPriceQtyAdd to Cart
    R0627S500 units10,000 units/ml$65.00Add to Cart
    R0627L2,500 units10,000 units/ml$260.00Add to Cart
      
    Categories:
    Nicking Endonucleases,
    Restriction Endonucleases: N-O
    Applications:
    DNA Nicking

    Description

    Nt.AlwI is a nicking endonuclease that cleaves only one strand of DNA on a double-stranded DNA substrate. It is an engineered derivative of AlwI which catalyzes a single strand break 4 bases beyond the 3´ end of the recognition sequence on the top strand.

    Product Source

    An E. coli strain containing a chimeric gene encoding the DNA recognition domain of AlwI and the cleavage/dimerization domain of Nt.BstNBI.

    Reagents Supplied

    The following reagents are supplied with this product:

    Store at (°C)Concentration
    CutSmart™ Buffer-2010X

    Properties and Usage

    Unit Definition

    One unit is defined as the amount of enzyme required to convert 1 µg of supercoiled pUC101 DNA (dam-/dcm-) to open circular form in 1 hour at 37°C in a total reaction volume of 50 µl.

    Reaction Conditions

    1X CutSmart™ Buffer
    Incubate at 37°C

    1X CutSmart™ Buffer:
    50 mM Potassium Acetate
    20 mM Tris-acetate
    10 mM Magnesium Acetate
    100 μg/ml BSA
    pH 7.9 @ 25°C

    Activity in NEBuffers2

    NEBuffer 1.1: 10%
    NEBuffer 2.1: 100%
    NEBuffer 3.1: 100%
    CutSmart™ Buffer: 100%

    Diluent Compatibility

    Storage Temperature

    -20°C

    Storage Conditions

    10 mM Tris-HCl
    50 mM KCl
    1 mM DTT
    0.1 mM EDTA
    200 μg/ml BSA
    50% Glycerol
    pH 7.4 @ 25°C

    Heat Inactivation

    80°C for 20 min

    Methylation Sensitivity

    dam methylation: Blocked
    dcm methylation: Not Sensitive
    CpG Methylation: Not Sensitive

    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.
    • Exonuclease Activity (Radioactivity Release):
      The product is tested in a reaction containing a radiolabeled mixture of single and double-stranded DNA. After incubation for 4 hours the exonuclease activity is determined by the % release of radioactive nucleotides.
    • Non-Specific DNase Activity (16 hour):
      The product is tested for non-specific nuclease degradation in a reaction containing a DNA substrate. After incubation for 16 hours there is no detectable degradation of the DNA substrate as determined by agarose gel electrophoresis.

    References

    1. Song, Q. et al. (2010). Anal. Chem. [Epub ahead of print].
    2. Zhang, P. et al. (2010). Protein Expr. Purif. 69, 226-234. [Epub 2009 Sep 9].

    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. My enzyme used to come with another NEBuffer, but CutSmart™ Buffer is now recommended?  Why?
    2. What effect does BSA have on the performance of NEB’s restriction enzymes when included in the new buffers?
    3. How can I access the old NEBuffer Activity Chart?
    4. How can I access the old Double Digest Finder?
    1. Optimizing Restriction Endonuclease Reactions
    2. Double Digest Protocol with Standard Restriction Enzymes

    Selection Tools

    Usage Guidelines & Tips

    Troubleshooting Guides

    Interactive Tools