Figure 4.

Quantification of the Genome-Editing Activity by Using the Relative Luciferase Activity Measurement

(A) Luciferase expression was verified by western blot with a luciferase-specific antibody. Only in lane 3 with gRNA, template, and Cas9 protein expression, a light luciferase band was visible. (B) RLU measured as a function of different combinations of gRNAs, Cas9 expression plasmid, and PAMin template DNA. Controls lack one of the necessary plasmids. (C) Comparison of PAMin versus PAMout template plasmid. The PAMout template plasmid results in 100% higher luciferase signal. (D) Influence of the DSB introduction either upstream, downstream, or at both sides of the template plasmid. The DSB induced by g3 RNA is more important than the one induced by the g4 RNA. All experiments were performed at least in triplicates. Error bars indicate the SD of the data set.