(A) The layout of the SpCas9 protein from C- (left) to N-terminus (right). Featuring the REC lobe (gray) and the domains of the NUC lobe: RuvC (light blue), BH (green), HNH (yellow), Topo-homology (Topo, pink), CTD (C-terminal domain, red). The last residue position for each domain is given below the domains. (B) Full structure (PDB: 4UN3 [46]) of SpCas9 in complex with sgRNA (orange), target DNA strand (dark blue), and displaced DNA strand (black, but barely visible here). Protein domains are colored in accordance with (A). Visualized using PyMOL [109]. (C) Same as B with some residues from the REC lobe removed to allow a view of the sgRNA and the target DNA strand. (D) Schematic overview of the R-loop formed by Cas9 RNP. DNA strands are colored in accordance with (B). The crRNA (orange) and tracrRNA (green) are shown connected by a linker (dashed lines between them) to form a sgRNA. The direction of R-loop formation is indicated with a dashed arrow. Approximate cleavage positions are represented by colored triangles.