Figure 7.

Optimization of guide RNA design and Cas9 delivery method. Imprecise repair outcomes in isogenic sex-1 strains differing only by a single base-pair change enabled comparison of the effectiveness of four RNA guides differing only in the penultimate nucleotide at the 3′ end of the target-specific sequence. Effectiveness of two different Cas9/guide delivery methods was compared for each guide: RNPs vs. DNA expression vectors. Using the DNA expression vector delivery approach, only the GG guide induced imprecise repair. Cas9 RNPs were more permissive, but the GG guide was still more effective than the other guides. Percentages represent the frequencies of imprecise repair events relative to the total number of dpy-10 co-converted F₁s (n).