DescriptionSacI has a High Fidelity version SacI-HF® (NEB #R3156).
High 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 minutes 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.
Product SourceAn E. coli strain that carries the SacI gene from Streptomyces achromogenes (ATCC 12767).
The following reagents are supplied with this product:
|Store at (°C)||Concentration|
Properties and Usage
Unit DefinitionOne unit is defined as the amount of enzyme required to digest 1 µg of λ DNA (HindIII digest) in 1 hour at 37°C in a total reaction volume of 50 µl.
1X NEBuffer 1.1
Incubate at 37°C
1X NEBuffer 1.1:
10 mM Bis-Tris-Propane-HCl
10 mM MgCl2
100 μg/ml BSA
pH 7 @ 25°C
Activity in NEBuffersNEBuffer 1.1: 100%
NEBuffer 2.1: 50%
NEBuffer 3.1: 10%
CutSmart® Buffer: 100%
10 mM Tris-HCl
100 mM NaCl
1 mM DTT
0.1 mM EDTA
200 μg/ml BSA
pH 7.4 @ 25°C
Heat Inactivation65°C for 20 min
dam methylation: Not Sensitive
dcm methylation: Not Sensitive
CpG Methylation: Not Sensitive
Quality Control AssaysThe 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. Further information regarding NEB product quality can be found here.
- Blue-White Screening (Terminal Integrity):
A sample of DNA vector linearized with a 10-fold excess of a restriction endonuclease, religated and transformed into an E. coli strain expressing the LacZ beta fragment gene results in less than 1% white colonies.
- Endonuclease Activity (Nicking):
The product is tested in a reaction containing a supercoiled DNA substrate. After incubation for 4 hours the percent converted to the nicked form is determined by agarose gel electrophoresis.
- 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.
This 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@example.com.
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.
- SacI is inhibited by salt concentrations > 10 mM. Mini-prep DNA containing residual salt is resistant to cleavage. A 70% alcohol wash or dialysis can be used to remove the salt.
- This enzyme has shown to have lower activity on some supercoiled plasmids, with more than 1 unit required to digest 1 μg plasmid DNA. For complete digestion of 1 μg of plasmid DNA please follow our recommended digestion protocol.
- Why isn't SacI cutting?
- How can the efficiency of SacI be increased?
- Is there any difference in the methylation sensitivity between SacI-HF and SacI?
- Is SacI affected by methylation?
- What is the molecular weight of SacI?
- What is the activity of SacI at 25°C?
- 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?
- 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?
- Alphabetized List of Recognition Specificities
- Average Fragment Size Generated By Endonuclease Cleavage
- Buffer and Diluent Formulation Table
- Compatible Cohesive Ends and Generation of New Restriction Sites
- Cross Index of Recognition Sequences
- Dam-Dcm and CpG Methylation
- Frequencies of Restriction Sites
- Time-Saver™ Qualified Enzymes
- Why Choose Recombinant Enzymes?
Usage Guidelines & 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
- Cleavage of Supercoiled DNA
- 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
- Reduced Star Activities of HF® Enzymes
- Restriction Endonucleases - Survival in a Reaction
- Restriction Enzyme Diluent Buffer Compatibility
- Restriction Enzyme Tips
- Single Letter Codes
- Star Activity
- Traditional Cloning Quick Guide