GenCrispr NLS-Cas9-EGFP is a fusion protein, containsa nuclear localisation signal (NLS) on its N terminal end, and an EGFP and a 6x(His) sequence on the C terminal end. Cas9 nuclease is an RNA-guided endonuclease that can catalyse cleavage of double stranded DNA. This kind of targeted nuclease is a powerful tool for genome editing with high precision.
- DNA-free: no external DNA added to system
- High cleavage efficiency: NLS ensures the efficient entry of the Cas9 protein into nuclei
- Low off target effects: transient expression of Cas9 nuclease improves specificity of cleavage
- Time-saving: no need for transcription and translation
- Reduced labour: enrich cell populations for desired genome edits via EGFP-based FACS, the C-terminal His-tag increases the choice of detection methods for the fusion protein
Cas9 protein forms a highly stable ribonucleoprotein (RNP) complex with the guide RNA (gRNA) component of the CRISPR/Cas9 system. When Cas9 is expressed with an NLS sequence, the Cas9 RNP complex can localise to the nucleus immediately upon entering the cell. There is no requirement for in vivo transcription or translation, which improves the efficiency of this method over plasmid-based systems. Additionally, the Cas9 RNP complex is rapidly cleared from the cells minimising the chance of off-target cleavage when compared to other systems.
Applications include screening for highly efficient and specific targeting gRNAs by in vitro DNA cleavage, in vivo gene editing when combined with a specific gRNA by electroporation or injection and enrichment of cell populations for desired genome edits via EGFP-based FACS.
Delivery information: GenCrispr NLS-Cas9-EGFP is supplied with 1X storage buffer (10 mM Tris, 300 mM NaCl, 0,1 mM EDTA, 1 mM DTT, 50% Glycerol pH7,4 at 25 °C).
