SfiI

Description

SfiI crystals (Lydia Dorner and Ira Schildkraut, New England Biolabs)

SfiI crystals (Lydia Dorner and Ira Schildkraut, New England Biolabs)

Product Source

An E. coli strain that carries the SfiI gene from Streptomyces fimbriatus (ATCC 15051).

Reagents Supplied

The following reagents are supplied with this product:

Store at (°C)Concentration
CutSmart® Buffer-2010X

Properties and Usage

Unit Definition

One unit is defined as the amount of enzyme required to digest 1 µg of Adeno-2 DNA in 1 hour at 50°C in a total reaction volume of 50 µl.

Reaction Conditions

1X CutSmart® Buffer
Incubate at 50°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 NEBuffers

NEBuffer 1.1: 25%
NEBuffer 2.1: 100%
NEBuffer 3.1: 50%
CutSmart® Buffer: 100%

Diluent Compatibility

Storage Temperature

-20°C

Storage Conditions

10 mM Tris-HCl
250 mM NaCl
0.1 mM EDTA
200 μg/ml BSA
50% Glycerol
0.15% Triton® X-100
1 mM DTT
pH 7.4 @ 25°C

Heat Inactivation

No

Methylation Sensitivity

dam methylation: Not Sensitive
dcm methylation: Impaired by Overlapping
CpG Methylation: Blocked by Some Combinations of Overlapping

Activity at Temperature

@37°C: 10%

Notes

  1. SfiI requires two copies of its recognition sequence for cleavage to occur. The two sites can be on either the same or different DNA molecules. Wertzell L.M. et al., (1995) J. Mol. Biol. 248:581-595.

    For more information and recommendations on working with enzymes requiring multi-sites, please visit Restriction Enzyme Cleavage: ‘single-site’ enzymes and ‘multi-site’ enzymes
  2. 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.

FAQs

  1. What is the minimum distance needed between two SfiI sites? Can they follow each other directly?
  2. Can I achieve digestion of a plasmid with a single SfiI site?
  3. Do degenerate recognition sites need to be palindromic?
  4. How can this enzyme be inactivated?
  5. Why isn't the enzyme cutting completely?
  6. Can I add an oligo to my reaction so that two sites are available for efficient cutting?
  7. What effect does BSA have on the performance of NEB’s restriction enzymes when included in the new buffers?
  8. Do I have to set-up digests with Time-Saver™ qualified enzymes for 5-15 minutes? Can I digest longer?
  9. How can I access the old NEBuffer Activity Chart?
  10. I tested your restriction enzyme on the substrate DNA recommended by NEB, and it appears to be active, however it does not digest my DNA. What could be the reason?
  11. Why do I see additional DNA bands on my gel after a restriction digest?
  12. Why is my Restriction Enzyme not cutting DNA?
  13. Why do I see a DNA smear on an agarose gel after a restriction digest?
  14. How many nucleotides do I have to add adjacent to the RE recognition site in order to get efficient cutting?

Protocols

  1. Optimizing Restriction Endonuclease Reactions
  2. Double Digest Protocol with Standard Restriction Enzymes
  3. Time-Saver Protocol for Restriction Enzyme Digests

Selection Charts

Feature Articles

Usage Guidelines & Tips

Troubleshooting Guides

Interactive Tools

Quality Control

Quality Control Assays

The 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.
  • 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.

Certificate of Analysis

The 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]

Specifications

The 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]

Safety Data Sheet

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.