Having supplied restriction enzymes to the research community for over 40 years, NEB has earned the reputation of being the leader in enzyme technologies. Working continuously to be worth of that distinction, NEB strives to develop enzyme of the highest purity and unparalleled quality.
Reduce Star Activity with High-Fidelity Restriction Enzymes
Time-Saver Protocol for Restriction Enzyme Digests
NEB TV Episode 15
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
Product SourceAn E. coli strain that carries the cloned TfiI gene from Thermus filiformis (D. Cowan, University College London).
The following reagents are supplied with this product:
|Store at (°C)||Concentration|
|CutSmart® Buffer||-20||10 X|
Properties and Usage
Unit DefinitionOne unit is defined as the amount of enzyme required to digest 1 µg of λ DNA in 1 hour at 65°C in a total reaction volume of 50 µl.
1X CutSmart® Buffer
Incubate at 65°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 NEBuffersCutSmart® Buffer: 100%
NEBuffer 1.1: 50%
NEBuffer 2.1: 100%
NEBuffer 3.1: 100%
10 mM Tris-HCl
250 mM NaCl
1 mM DTT
0.1 mM EDTA
200 μg/ml BSA
0.15% Triton® X-100
pH 7.4 @ 25°C
dam methylation: Not Sensitive
dcm methylation: Not Sensitive
CpG Methylation: Blocked by Some Combinations of Overlapping
Activity at Temperature@37°C: 10%
- 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.
- Cleavage of mammalian genomic DNA is blocked by some combinations of overlapping CpG methylation.
Faqs & Tech Tips
- What is Star Activity and how can it be avoided?
- Do degenerate recognition sites need to be palindromic?
- How can this enzyme be inactivated?
- My restriction enzyme used to be available at a lower concentration. Why does it now come at a higher concentration of 10,000 u/ml?
- What effect does BSA have on the performance of NEB’s restriction enzymes when included in the new buffers?
- Do I have to set-up digests with Time-Saver™ qualified enzymes for 5-15 minutes? Can I digest longer?
- How can I access the old NEBuffer Activity Chart?
- My restriction enzyme used to work well in the old NEBuffer but the new Performance chart indicates it has lower activity even though the only difference is the addition of BSA and removal of DTT to the new buffers. Why?
- 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?
- Why is my Restriction Enzyme not cutting DNA?
- How many nucleotides do I have to add adjacent to the RE recognition site in order to get efficient cutting?
Protocols & Manuals
Other Tools & Resources
- Alphabetized List of Recognition Specificities
- Buffer and Diluent Formulation Table
- Compatible Cohesive Ends and Generation of New Restriction Sites
- Cross Index of Recognition Sequences
- Dam-Dcm and CpG Methylation
- Enzymes with Multiple Recognition Sequences
- Frequencies of Restriction Sites
- Interrupted Palindromes
- Time-Saver™ Qualified Enzymes
- Why Choose Recombinant Enzymes?
Usage Guildelines & Tips
- Activity at 37°C for Restriction Enzymes with Alternate Incubation Temperatures
- Activity of Restriction Enzymes in PCR Buffers
- Cleavage Close to the End of DNA Fragments
- Digestion of Agarose-Embedded DNA: Info for Specific Enzymes
- Double Digests
- Effects of CpG Methylation on Restriction Enzyme Cleavage
- Heat Inactivation
- Megabase Mapping
- NEBuffer Activity/Performance Chart with Restriction Enzymes
- Optimizing Restriction Endonuclease Reactions
- 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
Troubleshooting GuidesRestriction Enzyme Troubleshooting Guide
Quality & Safety
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 control's 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.
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 firstname.lastname@example.org.
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.