Product SourceAn E. coli strain that carries the HincII gene from Haemophilus influenzae Rc (ATCC 49699).
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 in 1 hour at 37°C in a total reaction volume of 50 µl.
1X NEBuffer 3.1
Incubate at 37°C
1X NEBuffer 3.1:
100 mM NaCl
50 mM Tris-HCl
10 mM MgCl2
100 μg/ml BSA
pH 7.9 @ 25°C
Activity in NEBuffersNEBuffer 1.1: 25%
NEBuffer 2.1: 100%
NEBuffer 3.1: 100%
CutSmart™ Buffer: 100%
10 mM Tris-HCl
200 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: Blocked by Some Combinations of Overlapping
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.
- 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.
- A 100-fold overdigestion of HincII in NEBuffer 4 produces star activity.
- Cleavage of mammalian genomic DNA is blocked by some combinations of overlapping CpG methylation.
- Do degenerate recognition sites need to be palindromic?
- 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?
- How can I access the old Double Digest Finder?
- Alphabetized List of Recognition Specificities
- 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
- 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 a Q5®, Taq or Phusion PCR Mix
- Alteration of Apparent Recognition Specificities Using Methylases
- 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
- Restriction Endonucleases - Survival in a Reaction
- Restriction Enzyme Diluent Buffer Compatibility
- Restriction Enzyme Tips
- Single Letter Codes
- Star Activity
- Traditional Cloning Quick Guide