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  • NlaIII

    Description

    Product Source

    An E. coli strain that carries the NlaIII gene from Neisseria lactamica (NRCC 2118).

    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 digest 1 µg of ΦX174 RF I DNA 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 NEBuffers

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

    Diluent Compatibility

    Storage Temperature

    -80°C

    Storage Conditions

    10 mM Tris-HCl
    200 mM KCl
    1 mM DTT
    0.1 mM EDTA
    500 μg/ml BSA
    50% Glycerol
    pH 7.5 @ 25°C

    Heat Inactivation

    65°C for 20 min

    Methylation Sensitivity

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

    Shipping Notes

    • Ships on dry ice

    Notes

    1. NaCI and KCI inhibit activity, (NH4)2 SO4 does not.
    2. Cleaves to leave a 3´ CATG extension which can be ligated to DNA fragments generated by SphI.
    3. NlaIII retains full activity for 3 months at -20°C.
    4. For long term storage, aliquot and store at -80°C. 

    FAQs

    1. Is NlaIII inhibited by salt?
    2. What are the storage conditions for NlaIII?
    3. Does NlaIII produce commonly used compatible ends?
    4. Is extended digestion of NlaIII recommended?
    5. What effect does BSA have on the performance of NEB’s restriction enzymes when included in the new buffers?
    6. Do I have to set-up digests with Time-Saver™ qualified enzymes for 5-15 minutes? Can I digest longer?
    7. How can I access the old NEBuffer Activity Chart?
    8. I tested your restriction enzyme on the substrate DNA recommended by NEB, and it appears to be active, however it does not digest my DNA. What could be the reason?
    9. Why is my Restriction Enzyme not cutting DNA?
    10. Why do I see a DNA smear on an agarose gel after a restriction digest?
    11. Why do I see additional DNA bands on my gel after a restriction digest?
    12. How many nucletotides do I have to add adjacent to the RE recognition site in order to get efficient cutting?

    Tech Tips

    Low activity in other buffers because NaCl and KCl inhibit activity. Store at -70 deg C for long term storage.

    Protocols

    1. Optimizing Restriction Endonuclease Reactions
    2. Double Digest Protocol with Standard Restriction Enzymes
    3. Time-Saver Protocol for Restriction Enzyme Digests
    4. Protocol for Direct Digestion of gDNA during droplet digital PCR (ddPCR)
    5. Protocol for Digestion Prior to droplet digital PCR (ddPCR)

    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.

    Selection Tools

    Usage Guidelines & Tips

    Troubleshooting Guides

    Interactive Tools

    Citations

    • Sexton T, Kurukuti S, Mitchell JA, Umlauf D, Nagano T, Fraser P (2012). Sensitive detection of chromatin coassociations using enhanced chromosome conformation capture on chip Nat Protoc. 7(7), 1335-50. PubMedID: 22722369, DOI: 10.1038/nprot.2012.071

    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.
    • Ligation and Recutting (Terminal Integrity):
      After an over-digestion of DNA with a restriction endonuclease the percentage of the DNA fragments ligated with T4 DNA ligase and the percentage that can be recut are determined by agarose gel electrophoresis.
    • 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.

    Certificate of Analysis

    The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality control's for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

    Specifications

    The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

    Safety Data Sheet

    The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

    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.