CRISPR-Cas13d-mediated knockdown of sgms2a and sgms2b. (A) Schematic illustration of CRISPR-Cas13d-kd experimental design. (B) Embryo phenotype at 2 dpf. Embryos were classified into four categories: normal, mild deformity, severe deformity, and dead. Representative images of each category are presented on the left side of the graph. The WT, the sgms2a kd, the sgms2b kd, and the sgms2a+b kd embryos were compared with the sham embryos (Fisher’s exact test and Bonferroni correction). (C) Number of living fish during the 7-d experiment. The percentage of living fish at 2 and 7 dpf (the number of fish at 2 or 7 dpf compared with the original number of the injected embryos at day 0) for each group has been indicated in parenthesis. At day 7, larvae were euthanized for bone and cartilage staining. (D-G) Expression of sgms2a and sgms2b mRNA (relative expression to rpl13a) at 3.5 h post fertilization (hpf) and 2 dpf in sham, sgms2a kd, sgms2b kd, and sgms2a+b kd embryos by RT-qPCR. Data are shown as scatter dot plots with mean ± SD. The results from three independent experiments were combined. The sgms2a kd, the sgms2b kd, and the sgms2a+b kd embryos were compared with the sham embryos (ordinary one-way ANOVA for multiple comparisons). p < .05 was considered statistically significant. Only significant p-values were presented.
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