Cas9 in close-up

The Masc gene product controls masculinization in the black cutworm, Agrotis ipsilon

“…The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system is one of the simple and efficient genome editing tools (Barrangou et al, 2007;Cho et al, 2013;Cong et al, 2013). The Cas9-sgRNA (single guide RNA) complex recognizes and cleaves DNA in a sequencespecific manner, and random mutations are introduced at the sites of these double-stranded breaks (DSBs) (Brouns et al, 2008;Garside & MacMillan, 2014). CRISPR/Cas9 has been widely applied to edit genomes in B. mori and several other lepidopteran species.…”

Engineering Sequence-Specific DNA Binding Proteins for Antiviral Gene Editing

“…Recent solving of the crystal structure of Cas9 has aided understanding of the mechanism of DNA cleavage [81]. Studies performed on Cas9 from S. pyogenes in association with target and guide RNA revealed that the protein comprises two lobes with nuclease and target recognition (REC) functions [82].…”

An inducible and reversible genetics platform for novel drug target discovery

“…First, the target DNA must contain a "NGG" PAM. This is necessary because interaction between PAM and the phosphate lock on Cas9 pries apart the two DNA strands so that sgRNA can invade and form a Rloop with the DNA 114,120,121 . Once this happens, the second requirement comes into play: the 20 nt variable region of the sgRNA must match the target DNA sequence.…”

“…If there is only partial complementarity between the sgRNA and target DNA, cleavage will not occur [61][62][63] . If there is high complementarity, Cas9 undergoes a conformational change that puts the two nuclease lobes in position to nick both DNA strands 63,121 , creating a blunt end double-stranded break. The CRISPR/Cas9 system is a highly efficient technique for genetic engineering.…”