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Not sensitive to dam, dcm or mammalian CpG methylation.
High-Fidelity (HF®) restriction enzymes have the same specificity as native enzymes, but have been engineered for significantly reduced star activity and performance in a single buffer (CutSmart® Buffer). All HF-restriction enzymes come with Gel Loading Dye, Purple (6X). Enjoy the enhanced performance and added value of our engineered enzymes at the same price as the native enzyme:
- Engineered for improved performance
- 100% activity in CutSmart Buffer
- Time-Saver™ qualified for digestion in 5-15 minutes
- Reduced star activity
- Supplied with 1 vial of Gel Loading Dye, Purple (6X)
Featured VideosView Video Library
Reduce Star Activity with High-Fidelity Restriction Enzymes
Time-Saver Protocol for Restriction Enzyme Digests
NEB TV Ep. 15 – Applications of Restriction Enzymes
CutSmart™ Restriction Enzyme Buffer
Restriction Enzyme Digest Protocol: Cutting Close to DNA End
Restriction Enzyme Digestion Problem: DNA Smear on Agarose Gel
Why is My Restriction Enzyme Not Cutting DNA?
Restriction Enzyme Digest Problem: Too Many DNA Bands
Double Digestion with NEBcloner
DescriptionHigh Fidelity (HF) Restriction Enzymes have 100% activity in CutSmart Buffer; single-buffer simplicity means more straightforward and streamlined sample processing. HF enzymes also exhibit dramatically reduced star activity. HF enzymes are all Time-Saver qualified and can therefore cut substrate DNA in 5-15 with the flexibility to digest overnight without degradation to DNA. Engineered with performance in mind, HF restriction enzymes are fully active under a broader range of conditions, minimizing off-target products, while offering flexibility in experimental design.
NEB extensively performs quality controls on all standard and high-fidelity (HF) restriction enzymes. Examples of nuclease contamination studies for some of our HF restriction enzymes are shown below.
Product SourceAn E. coli strain that carries the cloned and modified BamHI gene from Bacillus amyloliquefaciens H (ATCC 49763)
The following reagents are supplied with this product:
|NEB #||Component Name||Component #||Stored at (°C)||Amount||Concentration|
- Product Categories:
- High-Fidelity (HF®) Restriction Endonucleases Products,
- Restriction Endonucleases B,
- time-saver-qualified-restriction-enzymes Products
- Restriction Enzyme Digestion
Properties & Usage
Unit DefinitionOne unit is defined as the amount of enzyme required to digest 1 ug of λ DNA in 1 hour at 37°C in a total reaction volume of 50 μl.
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 NEBuffersNEBuffer™ 1.1: 100%
NEBuffer™ 2.1: 50%
NEBuffer™ 3.1: 10%
CutSmart® Buffer: 100%
10 mM Tris-HCl
50 mM KCL
1 mM DTT
200 µg/ml BSA
0.1 mM EDTA
pH 7.4 @ 25°C
dam methylation: Not Sensitive
dcm methylation: Not Sensitive
CpG Methylation: Not Sensitive
- BamHI-HF® has the same specificity as BamHI (NEB #R0136), but it was engineered for reduced star activity.
- The increased specificity for the BamHI-HF® cut site has increased binding of the enzyme to the DNA. Under enzyme excess conditions the enzyme can remain attached to the DNA during gel electrophoresis. To disrupt binding, please use the supplied 6X Purple Gel Loading Dye (NEB #B7024S) or add SDS to a final concentration of 0.1% - 0.5%. Alternatively to purify the DNA we recommend Monarch PCR & DNA Cleanup kit, NEB #T1030) before electrophoresis.
- For enzymes that cannot be heat-inactivated, we recommend using a column for cleanup (such as the Monarch® PCR & DNA Cleanup Kit), or running the reaction on an agarose gel and then extracting the DNA (we recommend Monarch Gel Extraction Kit), or performing a phenol/chloroform extraction.
- Not sensitive to CpG, dcm, or dam methylation.
Protocols, Manuals & Usage
Usage & Guidelines
- Activity at 37°C for Restriction Enzymes with Alternate Incubation Temperatures
- Activity of Restriction Enzymes in PCR Buffers
- Alteration of Apparent Recognition Specificities Using Methylases
- Cleavage Close to the End of DNA Fragments
- Dam and Dcm Methylases of E. coli
- Digestion of Agarose-Embedded DNA: Info for Specific Enzymes
- Double Digests
- Effects of CpG Methylation on Restriction Enzyme Cleavage
- Heat Inactivation
- NEBuffer Activity/Performance Chart with Restriction Enzymes
- Optimizing Restriction Endonuclease Reactions
- Reduced Star Activities of HF® Enzymes
- Restriction Endonucleases - Survival in a Reaction
- Restriction Enzyme Diluent Buffer Compatibility
- Restriction Enzyme Tips
- Restriction Enzymes for Droplet Digital PCR (ddPCR)
- Single Letter Codes
- Star Activity
- Traditional Cloning Quick Guide
Tools & Resources
- Alphabetized List of Recognition Specificities
- Cleavage of Supercoiled DNA
- Compatible Cohesive Ends and Generation of New Restriction Sites
- Cross Index of Recognition Sequences
- Dam-Dcm and CpG Methylation
- Frequencies of Restriction Sites
- Recleavable Filled-in 5' Overhangs
- Time-Saver™ Qualified Enzymes
- Why Choose Recombinant Enzymes?
FAQs & Troubleshooting
- Is there a difference in cutting close to the ends between BamHI-HF and BamHI?
- Is there any difference in the methylation sensitivity between BamHI-HF and BamHI?
- What is the difference between BamHI-HF and BamHI?
- How does the level of star activity of BamHI-HF compare to BamHI?
- Why are there larger than expected bands after digestion with the BamHI-HF™ restriction enzyme?
- What does HF® refer to following the name of a restriction enzyme?
- When is star activity a concern?
- Why does the HF version of the enzyme have a different recommended buffer than the wild type enzyme?
- When should I choose the High Fidelity (HF®) version of the enzyme?
- How is the improvement in fidelity of HF restriction endonucleases quantitated?
- What is the Fidelity Index (FI)?
- Can the change in buffer preference of the HF enzyme be advantageous?
- Will the HF enzyme produce elevated star activity when used in a buffer other than the one recommended?
- What does it mean to be Time-Saver™ qualified?
- Do I have to set-up digests with Time-Saver™ qualified enzymes for 5-15 minutes? Can I digest longer?
- 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?
- Is Gel Loading Dye, Purple (6X) or Gel Loading Dye, Purple (6X), no SDS compatible with other DNA binding dyes such as SYBR® and GelRed™ during gel electrophoresis?
- Can Gel Loading Dye, Purple 6X (B7024) be stored in cold temperatures?
- Why is my Restriction Enzyme not cutting DNA?
- Why do I see additional DNA bands on my gel after a restriction digest?
- Why do I see a DNA smear on an agarose gel after a restriction digest?
- How many nucleotides do I have to add adjacent to the RE recognition site in order to get efficient cutting?
- Which NEB restriction enzymes are supplied with Gel Loading Dye, Purple (6X)?
- What is the specific activity of BamHI-HF?
Quality, Safety & Legal
Quality Assurance StatementQuality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.
SpecificationsThe 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]
Certificate Of AnalysisThe Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]
Safety DataSheetsThe following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.
Gel Loading Dye, Purple (6X)
Legal and DisclaimersThis product is covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB).
While NEB develops and validates its products for various applications, the use of this product may require the buyer to obtain additional third party intellectual property rights for certain applications.
For more information about commercial rights, please contact NEB's Global Business Development team at [email protected].
This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.
The supporting documents available for this product can be downloaded below.