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  • PhoI

    The product PhoI (R0705S/L) was discontinued effective August 6, 2013. The isoschizomer HaeIII (R0108S/L) is a suitable alternative for most applications.
    • This product was discontinued on 08/09/2013
    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 no cpg dcm
    Pho-I-cutsite_1
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    Categories:
    Discontinued Products

    Description

    Product Source

    An E. coli strain that carries the PhoI gene from Pyrococcos horikoshii OT3 (Y. Kawarabayasi).

    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 λ DNA in 1 hour at 75°C in a total reaction volume of 50 µl.

    Reaction Conditions

    1X CutSmart® Buffer
    Incubate at 75°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: 50%
    NEBuffer 2.1: 50%
    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

    No

    Methylation Sensitivity

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

    Activity at Temperature

    @37°C: 0%

    Notes

    1. PhoI is a highly thermostable restriction enzyme that can survive temperatures as high as 95°C. Please inquire about other enzymes in this class that are in development at NEB.

    FAQs

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

    Protocols

    1. Optimizing Restriction Endonuclease Reactions
    2. Double Digest Protocol with Standard Restriction Enzymes

    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

    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.

    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.