Product SourceAn E. coli strain that carries the cloned StyD4I gene from Salmonella typhi D4 (E.S. Anderson).
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 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: 10%
NEBuffer 2.1: 100%
NEBuffer 3.1: 100%
CutSmart® Buffer: 100%
10 mM Tris-HCl
300 mM NaCl
1 mM DTT
0.1 mM EDTA
500 μg/ml BSA
pH 7.4 @ 25°C
Heat Inactivation65°C for 20 min
dam methylation: Not Sensitive
dcm methylation: Blocked by Overlapping
CpG Methylation: Impaired by 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.
- StyD4I is a neoschizomer of ScrFI.
- Blocked by overlapping dcm methylation. Cleavage of mammalian genomic DNA is impaired by overlapping CpG methylation.
- StyD4I (CCNGG) has 4 possible recognition sequences: two, CCAGG and CCTGG can be blocked by dcm methylation CCWGG (W = A or T). The other two recognition sites CCCGG and CCGGG are not blocked by dcm methylation. Note dcm methylation can be removed or avoided by using the NEB dam-/dcm- Competent E. coli strain (NEB #C2925).
- Do degenerate recognition sites need to be palindromic?
- When is StyD4I blocked by overlapping dcm methylation?
- My enzyme used to come with another NEBuffer, but CutSmart™ Buffer is now recommended? Why?
- 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?