Fig. 1

Analysis of DSB repair patterns in CRISPR/Cas9-mediated embryo editing. A Schematic representation of the analysis of DSB repair patterns in CRISPR/Cas9-mediated embryo gene editing. B A summary of the editing efficiency for the tested sgRNAs. The blue scale represents the editing efficiency of insertions and deletions, while the green scale illustrates the overall editing efficiency. C CRISPR-Cas9 cleavage and its subsequent repair processes are depicted, where the PAM sequence is denoted by red bases, the insertion sequence by purple bases, the deletion sequence by a yellow dotted line, and the micro-homologous arm by a red underline. D The repair profile of Calcr-sgRNA2 is presented, wherein the PAM sequence (NGG) is highlighted in red, the missing sequence is represented by a gray base region, and the micro-homologous sequence is underlined in red. E The relationship between sgRNA editing efficiency and the NHEJ/MMEJ ratio. Each circle corresponds to a specific sgRNA, with Calcr-sg2 highlighted. Data are presented as mean ± standard error of the mean (s.e.m.). F The relative expression levels of the indicated genes (Rad52, Ku70, and Polq). The “Blank” group refers to embryos without any treatment, the “Dilution solvent” group corresponds to embryos injected with dilution buffer, the “Cas9 only” group represents embryos injected with Cas9 mRNA only, and the “Donor only” group signifies embryos injected with donor DNA only. A two-sided Student’s t-test was used for statistical analysis, with *p < 0.05 and **p < 0.01 considered significant. Data are presented as mean ± standard error of the mean (s.e.m.). Each bar includes at least two biological replicates. G Gene knockdown via CasRX. RT-PCR analysis revealed the expression levels of Rad52, Ku70, and Polq in both control and CasRX-treated embryos. The orange bar denotes the Cas9/scramble gRNA group, which serves as a control. A two-sided Student’s t-test was used for statistical analysis, with *p < 0.05 and ***p < 0.001 considered significant. Data are presented as mean ± standard error of the mean (s.e.m.). Each bar includes at least three biological replicates