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EnGen® Spy Cas9 Nickase

Catalog # Concentration Size List Price Quantity Your Price
M0650S 1 µM 90 pmol $91.00
$81.90
M0650T 20 µM 500 pmol $229.00
$206.10
Catalog # Size List Price Your Price
M0650S 90 pmol $91.00
$81.90
M0650T 500 pmol $229.00
$206.10
Catalog #
Qty:
 
*On-line ordering is for Canadian customers only. Web pricing is applicable only to orders placed online at www.neb.ca
  • Variant of Cas9 nuclease differing by a point mutation (D10A) in the RuvC nuclease domain
  • Capable of generating nicks, but not cleaving DNA
  • DNA double strand breaks can be generated, with reduced off-target cleavage, by targeting two sites with EnGen Cas9 Nickase in close proximity
EnGen® Spy Cas9 Nickase is a variant of Cas9 nuclease differing by a point mutation (D10A) in the RuvC nuclease domain, which enables it to nick, but not cleave, DNA (1,2). EnGen Spy Cas9 Nickase, like EnGen Cas9 NLS (NEB #M0646), targets DNA using a guide RNA complementary to a site with a 3’ NGG protospacer adjacent motif (PAM). Nicking occurs on the DNA strand opposite the PAM. Double-stranded DNA breaks can be generated with reduced off-target cleavage by targeting two sites in close proximity (generally 0-20 bp apart) and with PAMs facing outward to leave 5’ overhangs. Nicking by EnGen Spy Cas9 Nickase occurs on the DNA strand opposite the target sequence. EnGen Spy Cas9 Nickase contains SV40 T antigen nuclear localization (NLS) sequences on the N and C- termini.
Product Source
EnGen® Spy Cas9 Nickase is expressed as a N-terminal 6XHis-tagged fusion in E. coli.
Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  EnGen® Spy Cas9 Nickase M0650SVIAL -20 1 x 90 pmol 1 µM
  NEBuffer™ r3.1 B6003SVIAL -20 1 x 1.25 ml 10 X
  EnGen® Spy Cas9 Nickase M0650TVIAL -20 1 x 500 pmol 20 µM
  NEBuffer™ r3.1 B6003SVIAL -20 1 x 1.25 ml 10 X
Application Features
  • Paired, offset cleavage of genomic loci
  • Target-specific DNA nicking

Properties & Usage

Reaction Conditions

1X NEBuffer™ r3.1
Incubate at 37°C

1X NEBuffer™ r3.1
100 mM NaCl
50 mM Tris-HCl
10 mM MgCl2
100 µg/ml Recombinant Albumin
(pH 7.9 @ 25°C)

Storage Buffer

300 mM NaCl
10 mM Tris-HCl
0.1 mM EDTA
1 mM DTT
50% Glycerol
pH 7.4 @ 25°C

Heat Inactivation

65°C for 5 minutes

Notes
  • 20 µM is equal to ~3.2 mg/ml EnGen® Cas9 Nickase NLS, S. pyogenes. 1 µM is equal to ~0.16 mg/ml.
  • This product is compatible with Diluent Buffer B: 300 mM NaCl, 10 mM Tris-HCl, 0.1 mM EDTA, 1 mM DTT, 500 µg/ml BSA and 50% glycerol. (pH 7.4 @ 25°C).
References
  • Mali, P., et. al. (2013). NatBiotech. 31 (9), 838-8.
  • Ran, FA., et. al. (2013). Cell. 154 (6), 1380-9.
Quality Control Assay
Quality 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.
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]
Certificate of Analysis
The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]
Safety Data Sheets
Legal And Disclaimer

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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.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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