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  • BcgI

    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 sam cpg dam
    Bcg-I-cutsite_1
    Catalog #SizeConcentrationPriceQtyAdd to Cart
    R0545S250 units2,000 units/ml$63.00Add to Cart
    R0545L1,250 units2,000 units/ml$252.00Add to Cart
      
    Categories:
    Restriction Endonucleases: B
    Applications:
    Restriction Enzyme Digestion

    Description

    Product Source

    An E. coli strain that carries the BcgI gene from Bacillus coagulans (H. Kong).

    Reagents Supplied

    The following reagents are supplied with this product:

    Store at (°C)Concentration
    S-adenosylmethionine (SAM)-2032 mM
    NEBuffer 3.1-2010X

    Properties and Usage

    Unit Definition

    One unit is defined as the amount of enzyme required to digest 1 µg of λ DNA in 1 hour at 37°C in a total reaction volume of 50 µl.

    Reaction Conditions

    1X NEBuffer 3.1
    Supplement with 20 μM S-adenosylmethionine (SAM)
    Incubate at 37°C

    1X NEBuffer 3.1:
    100 mM NaCl
    50 mM Tris-HCl
    10 mM MgCl2
    100 μg/ml BSA
    pH 7.9 @ 25°C

    Activity in NEBuffers

    NEBuffer 1.1: 10%
    NEBuffer 2.1: 75%
    NEBuffer 3.1: 100%
    CutSmart™ Buffer: 50%

    Diluent Compatibility

    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

    65°C for 20 min

    Methylation Sensitivity

    dam methylation: Impaired by Overlapping
    dcm methylation: Not Sensitive
    CpG Methylation: Blocked by Some Combinations of Overlapping

    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.

    Notes

    1. BcgI cleaves DNA substrates twice to excise its recognition site generating a 32 base-pair fragment with 2-base 3´ overhangs.
    2. Requires S-adenosylmethionine for optimal activity (supplied with enzyme). S-adenosylmethionine is stored at -20°C as a 32 mM solution dissolved in sulfuric acid (0.005 M) and 10% ethanol. SAM in this solution stored under ideal conditions remains active for up to 6 months. SAM is unstable at pH 7.5 and 37°C and should be replenished for reactions incubated longer than 4 hours. Many problems in achieving complete digestion can be alleviated by using fresh SAM.
    3. May exhibit star activity in NEBuffer 2.1, or CutSmart Buffer.

    Supporting Documents

    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]

    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. Why isn't BcgI cutting my DNA?
    2. Does BcgI require SAM for activity?
    3. Do BcgI generated ends ligate?
    4. What effect does BSA have on the performance of NEB’s restriction enzymes when included in the new buffers?
    5. How can I access the old NEBuffer Activity Chart?
    6. How can I access the old Double Digest Finder?
    1. Optimizing Restriction Endonuclease Reactions

    Selection Tools

    Usage Guidelines & Tips

    Troubleshooting Guides

    Interactive Tools