SpCas9 (D10A, H840A)-NLS, Active(CS09N-E333H)

SpCas9 (D10A, H840A)-NLS, Active(CS09N-E333H)

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Description :Recombinant full length SpCas9 (D10A, H840A)-NLS from S. pyogenes was expressed in E. coli using a C-terminal His tag.

Species :Streptococcus pyogenes

Tag :HIS

Expression System:E.coli

Sequence :Full length

Genbank Number :AAK33936

Alternative name(s) :Cas9 (D10A, H840A)-Nikase Protein. UniProt accession Q99ZW2

Formulation :Recombinant protein stored in 50mM sodium phosphate, pH 7.5, 300mM NaCl, 200mM imidazole, 1mM DTT, 10% glycerol.

Storage and Stability :Store product at –70oC. For optimal storage, aliquot target into smaller quantities after centrifugation and store at recommended temperature. For most favorable performance, avoid repeated handling and multiple freeze/thaw cycles. Always include RNase inhibitor at manufacturer recommended concentrations in Cas enzyme reactions.

Scientific Background :Cas9 is a multidomain nuclease from the bacterium Streptococcus pyogenes characteristic of type II CRISPR- Cas systems (1). Using single guide RNA (sgRNA), Cas9 is targeted to specific genomic loci where it generates a blunt ended double strand beak (2). SpCas9 (D10A, H840A)-NLS is a catalytic mutant of Cas9 containing a nuclear localization sequence for optimal activity. Unlike wild type Cas9, SpCas9 (D10A, H840A)-NLS generates staggered double strand DNA breaks with overhangs (3). SpCas9 (D10A, H840A) can be used for various genome editing applications.

References :1. Nishimasu H, Ran FA, Hsu PD, Konermann S, Shehata SI, Dohmae N, Ishitani R, Zhang F, Nureki O. Crystal structure of Cas9 in complex with guide RNA and target DNA. 2014. Cell. 156(5):935-49.
2. Sapranauskas R, Gasiunas G, Fremaux C, Barrangou R, Horvath P, Siksnys V. 2011. The Streptococcus thermophilus CRISPR/Cas system provides immunity in Escherichia coli. Nucleic Acids Res. 39(21):9275-82.
3. Jiang F, Doudna JA. 2017. CRISPR-Cas9 Structures and Mechanisms. Annu Rev Biophys.46:505-529.

Purity :Sample Purity Data. For specific information on a given lot, see related technical data sheet.

Molecular Weight :Observed MW ~160 kDa Calculated MW ~161 kDa

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Research Areas :Cancer