Fig 1: Validation of an all-in-one AAV-sgRNA-hNmeCas9 construct. a Schematic representation of a single rAAV vector expressing human-codon-optimized NmeCas9 and its sgRNA. The backbone is flanked by AAV inverted terminal repeats (ITR). The poly(a) signal is from rabbit beta-globin (BGH). b Schematic diagram of the Pcsk9 (top) and Rosa26 (bottom) mouse genes. Red bars represent exons. Zoomed-in views show the protospacer sequence (red) whereas the NmeCas9 PAM sequence is highlighted in green. Double-stranded break location sites are denoted (black arrowheads). c Stacked histogram showing the percentage distribution of insertions-deletions (indels) obtained by TIDE after AAV-sgRNA-hNmeCas9 plasmid transfections in Hepa1–6 cells targeting Pcsk9 (sgPcsk9) and Rosa26 (sgRosa26) genes. Data are presented as mean values ± SD from three biological replicates. d Stacked histogram showing the percentage distribution of indels at Pcsk9 in the liver of C57Bl/6 mice obtained by TIDE after hydrodynamic injection of AAV-sgRNA-hNmeCas9 plasmid