EnGen® Spy Cas9 Nickase
- 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 SourceEnGen® Spy Cas9 Nickase is expressed as a N-terminal 6XHis-tagged fusion in E. coli.
The following reagents are supplied with this product:
Store at (°C) Concentration NEBuffer™ 3.1 -20 10X
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