Figure 2. Multiplex genome editing with Cas9 nuclease and seven gRNAs.

(A) Schematic illustration of the all-in-one vector expressing seven gRNAs targeting seven different genomic loci and Cas9 nuclease. The blue and green bent arrows indicate the U6 and CBh promoters, respectively. (B) Genomic cleavage analysis of the seven genomic loci targeted with the all-in-one CRISPR/Cas9-nuclease vector. The products from untransfected control cells (C) and cells transfected with CRISPR/Cas9-nuclease vectors targeting seven (7) and single (1) loci were analyzed by agarose gel electrophoresis. The percentage of non-homologous end-joining (% NHEJ) was estimated using ImageJ software as previously described³⁰.