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Recombinant full-length Streptococcus pyogenes Cas9 was expressed in E. coli with a C-terminal His tag.
CS09-E311H-50 |
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概述:
Streptococcus pyogenes Cas9 is a type II-A Cas9 protein and is the most used in most current CRISPR-based genetic engineering methodologies (1). CRIPSR RNAs (crRNAs) fused to trans-activating CRISPR RNAs (tracrRNA) direct sequence-specific cleavage by Cas9, introducing double-stranded breaks in cell-invading DNA in bacterial and archaeal CRIPSR/Cas9 systems (1,2). Cas9 requires the 5’-NGG-3’ protospacer adjacent motif (PAM) to destabilize the target DNA (3). Cas9’s HNH endonuclease domain cleaves the strand complementary to the crRNA while its RuvC endonuclease domain cleaves the non-complementary strand (2).
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参考文献:
1. Jinek, M. et al. Structures of Cas9 Endonucleases Reveal RNA-Mediated Conformational Activation. Science. 2014, 343(6176): 1247997.
2. Jinek, M. et al. A programmable dual RNA-guided endonuclease in adaptive bacterial immunity. Science. 2012, 337(6096): 816-821.
3. Anders, C. et al. Structural basis of PAM-dependent target DNA recognition by the Cas9 endonuclease. Nature. 2014, 513(7519): 569-573.
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Cancer, Cell Cycle