For over 40 years, New England Biolabs has been developing innovative solutions for molecular biology applications. The respected leader in the field of restriction enzyme biology, NEB has developed a line of High-Fidelity (HF®) Restriction Enzymes. These engineered enzymes have the same specificity as the native enzyme, with the added benefit of reduced star activity, rapid digestion (5-15 minutes) and 100% activity in CutSmart® Buffer.
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.
Optimized performance for a wide range of conditions
High Fidelity Restriction Enzymes have been engineered by exchanging functional amino acid residues and then screening for optimal mutants that perform under a wide range of conditions. Whether you are setting up digests for 5-15 minutes or overnight, or using varying amounts of enzymes, HF enzymes ensure the performance you need.
HF® is a registered trademark of New England Biolabs, Inc.
FastDigest® is a registered trademark of Fermentas.
FAQs for High-Fidelity (HF®) Restriction Endonucleases
Protocols for High-Fidelity (HF®) Restriction Endonucleases
- Digestion of Agarose-Embedded DNA
- Determining Genome Targeting Efficiency using T7 Endonuclease I
- Double Digest Protocol with Standard Restriction Enzymes
- Protocol for Direct Digestion of gDNA during droplet digital PCR (ddPCR)
- Protocol for Digestion Prior to droplet digital PCR (ddPCR)
- Optimizing Restriction Endonuclease Reactions
A Modern Day Gene Genie Sir Richard Roberts on Rebase
Restriction Enzymes at NEB: Over 30 years of Innovation
Restriction Endonucleases Technical Guide
The Restriction Enzyme Technical Guide provides product information and technical reference charts on the wide range of restriction enzymes available from NEB.
- DNA Sequences and Maps Tool
- Enzyme Finder
- Alphabetized List of Recognition Specificities
- Average Fragment Size Generated By Endonuclease Cleavage
- Compatible Cohesive Ends and Generation of New Restriction Sites
- Cross Index of Recognition Sequences
- Dam-Dcm and CpG Methylation
- Enzymes with Multiple Recognition Sequences
- Enzymes with Nonpalindromic Sequences
- Frequencies of Restriction Sites
- Interrupted Palindromes
- Recleavable Blunt Ends
- Recleavable Filled-in 5' Overhangs
- Restriction Enzyme Troubleshooting Guide
- 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
- Cleavage Of Supercoiled DNA
- Dam and Dcm Methylases of E. coli
- Digestion of Agarose-Embedded DNA: Info for Specific Enzymes
- Double Digests
- Heat Inactivation
- Megabase Mapping
- 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 of Foreign DNA by E. coli K-12
- Site Preferences
- Star Activity
Other Tools & Resources
- Kamps-Hughes, N., Quimby, A., Zhu, Z., Johnson, E.A. 2013. Massively parallel characterization of restriction endonucleases Scopus. 41(11), PubMedID: 23605040, DOI: 10.1093/nar/gkt257
Publications related to High-Fidelity (HF®) Restriction Endonucleases
Benefits of High-Fidelity (HF®) Restriction Enzymes
- One buffer convenience with no loss of performance
- Reduced star activity eliminates unwanted cleavage
- Time-Saver qualified for 5-15 minute digests, flexible enough to digest overnight.
- Engineered for performance under a wide range of conditions
- Added flexibility without added cost
- Mutational Analysis
- Probe Preparation
- Methylation Detection
- Any application that requires high fidelity or flexible reaction setup
NEB has engineered HF® enzymes to eliminate star activity. Learn how, and what this means for your digests.
Watch as Rick Morgan, Research Scientist in the Restriction Enzyme Division, describes his passion for discovering and characterizing restriction enzymes from nature.
Watch as Geoff Wilson, Restriction Enzyme Division Head, describes the interaction of restriction enzymes and substrate DNA using computer models generated from x-ray crystallography data.
Are you finding unexpected bands in your digestion reaction? Here are some tips to help you determine the cause.
Learn what Star Activity is, why it is detrimental to accurate restriction enzyme digestion, and how NEB's HF enzymes are engineered to avoid it.
Double digestions can save you time, and this video can offer tips for how to achieve the best results, no matter which of NEB's restriction enzymes you're using.
Learn more about what causes this common problem, and how NEB's enzymes are QC'd to avoid DNA smearing.
Let one of NEB's restriction enzyme experts help you improve your technique and avoid common mistakes in digest setup.
>210 of NEB's restriction enzymes are 100% active in a single buffer. Learn more about CutSmart® Buffer and why it matters to you.