Cas9 (D10A) Nickase, Nuclear Localization Sequence (NLS)

NLS-Cas9-D10A Nickase is a mutation form of Cas9 Nuclease. Cas9 nuclease is an RNA-guided endonuclease that can catalyze cleavage of double stranded DNA. This kind of targeted nuclease is a powerful tool for genome editing with high precision. Cas9 nuclease cleaves the double strand DNA generating two break sites based on its two active domains. NLS-Cas9-D10A Nickase is a mutation form of Cas9 Nuclease which makes one active domain deactivated, thus it can only cut one single strand DNA that is complementary to the guide-RNA, producing one single strand cut. Combined with two different gRNA, NLS-Cas9-D10A Nickase produces two cut sites respectively and causes a double strand break. Compared with the wild type Cas9 nuclease, the two-gRNA guided cleavage can significantly reduce the off target effects.

NLS-Cas9-D10A Nickase is highly purified mutant proteins expressed in an E. coli strain carrying a plasmid encoding the mutated Cas9 gene from Streptococcus pyogenes.

Applications:
Screening the highly efficient and specific targeting gRNAs using in vitro DNA cleavage. In vivo gene editing combined with specific gRNA by electroporation or injection.

Bioactivity:
>90% cleavage efficiencies

Endotoxin:
<0.1EU/ug

Key Features:
DNA-free: no external DNA added to system.
High cleavage efficiency: NLS ensures the efficient entry of Cas9 protein into nuclei.
Low off target: Double gRNA-guided cleavage and transient expression of Cas9 nuclease.
Time-saving: no need for transcription and translation.

Supplied With:
Reaction Buffer, 10X
Supplied as a liquid in 100mM Tris-HCl, 300mM sodium chloride, pH 7, 0.1mM EDTA, 1mM DTT, 50% glycerol.

Storage and Stability:
May be stored at 4°C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20°C. Aliquots are stable for 6 months after receipt at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.