Generation of med27 LoF zebrafish. A Diagram illustrating the microinjection of the CRISPR/Cas9 genome editing system into zebrafish embryos to introduce med27 null alleles. B Structure of the med27 gene showing the sgRNA targeting site and the resulting med27 frameshift mutations. E1 to E8: exons 1 to 8; PAM: protospacer adjacent motif; WT: wildtype; INS: insertion; DEL: deletion. C Representative lateral and dorsal views, along with quantification of med27 expression in WT (med27+/+), med27+/−, and med27−/− embryos, detected by WISH. Hybridization images were obtained using the med27 antisense probe, with quantification shown for 24 hpf embryos from the INS10 line. For WISH results at various time points in all three lines, refer to Fig. S2A. Control: WISH with the sense probe. Scale bar = 500 μm. D Expression of med27 in med27+/+, med27+/−, and med27−/− larvae from the INS10 line at 7 dpf as detected by RT-qPCR. Two sets of primers (med27-1, located upstream of the induced mutation site, and med27-2, null downstream of the mutation site) were used in RT-qPCR to quantify med27 mRNA. For RT-qPCR results in the INS5 and DEL5 lines, refer to Fig. S2B. Error bars represent mean ± standard deviation (SD). Statistical analysis was performed using one-way ANOVA. *: P<0.05; ***: P<0.001; ****: P<0.0001
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