Figure 3

Figure 3

The InDelphi-mESC model accurately predicts CRISPR/Cas9 gene editing outcomes in X. tropicalis, X. laevis and zebrafish embryos which can be exploited to identify high-frameshift frequency gRNAs. (A–F) Scatter plot with InDelphi-mESC-predicted cumulative frameshift gene editing frequencies correlated to experimentally observed cumulative frameshift gene editing frequencies, for each sgRNA separately, in X. tropicalis (n = 14) (Panel A), in X. laevis (n = 6) (Panel B) and in zebrafish (n = 15) embryos (Panel C). Scatter plot with InDelphi-mESC-predicted INDEL patterns correlated to experimentally observed INDEL patterns, for all gRNAs simultaneous, in X. tropicalis (n = 14) (Panel D), in X. laevis (n = 6) (Panel E) and zebrafish (n = 15) (Panel F) embryos. Black demarcated lines show the perfect correlation r = 1. Light-grey areas show the standard error on the best-fit linear regression line. Black lines show linear regression model. (G) Correlations between model-predicted INDEL patterns to experimentally observed INDEL patterns, for each gRNA separately. Correlations for X. tropicalis embryos (n = 14) (dark blue) and X. laevis embryos (n = 6) (middle blue) analyzed by Sanger sequencing and sequence trace decomposition. Correlations for zebrafish embryos analyzed by targeted amplicon sequencing (TAS) (n = 15) (light blue). (H) Using the distribution of the expected probability of frameshift frequency for a large dataset of SpCas9 human target sites in mESC cells from Shen et al. 2018 (black line—monoallelic)²⁷, we draw the derivative distribution of the probability of a randomly designed gRNA to generate biallelic frameshift editing. This distribution is shown for different editing efficiencies within the F₀ mosaic animal: 100%, 50% and 25% (in reducing intensities of blue—100 circles, each circle represents a cell within a total mosaic of a 100 cells). E.g. The probability of a randomly designed gRNA to yield more than 80% biallelic frameshift mutant cells in a developing mosaic, assuming 100% efficiency, is the area under curve highlighted in pink and represents only a 3.24% probability.