Fig. 3

Injection of dual gRNAs individually targeting the C. briggsae and the C. nigoni genome significantly increases the efficiency of targeted recombination. A Pie charts showing the ratio between alignable (blue) and unalignable (brown) sequences for the introns and intergenic regions of one-to-one ortholog pairs between C. briggsae and C. nigoni. B Violin plots with overlaid boxplots showing the similarity of alignable coding sequences (CDS), introns, or intergenic sequences of one-to-one ortholog pairs. (****) p < 0.0001 (Wilcoxon ranked sum test with multiple testing correction using FDR method). C Shown is a recombination resistant region with a relatively low sequence homology and two gRNAs each targeting C. briggsae (CBG16459) and C. nigoni (g17687), respectively. D Bar plots showing the comparison of targeted recombinant frequency between the regions with a relatively high (Fig. 2D) and low sequence homology (C) using C. briggsae-specific gRNAs. (****) p < 0.0001 (chi-square test). Number of screened crossing progeny (n) is indicated in parenthesis. E Comparison of targeted recombinant frequency using a single or dual gRNA(s). Left: schematic diagrams of double-stranded DNA breaks induced by a single or dual species-specific gRNA(s). Right: bar plots showing the comparison of recombinant frequency between the control (no gRNA), and the treatment with a single or dual species-specific gRNA(s). Note that the animals treated with dual gRNAs display a significantly higher targeted recombinant frequency. (****) p < 0.0001 (Fisher’s exact test with multiple testing correction using FDR method)