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Fig. 8 Downregulation of EZH2 disrupts the rhythmic expression of circadian clock genes in zebrafish Under LD and DD conditions, RNA samples were collected every 4 h from 5 dpf wild-type (WT) and ezh2−/− mutant zebrafish larvae (n = 30) for RT‒qPCR analysis. (A–D) Under LD conditions, RT‒qPCR was performed on the core clock genes (clock1a, bmal1b, per1b, per3) of 5 dpf wild-type (WT) and ezh2−/− mutant zebrafish larvae. The experiments were repeated three times. (E–H) RNA samples were collected from 30 wild-type (WT) and 5 dpf ezh2−/− mutant zebrafish larvae under DD conditions every 4 h. RT‒qPCR was conducted on core clock genes (clock1a, bmal1b, per1b, and per3), and the experiments were repeated three times. (I–M) Zebrafish embryos were microinjected with morphine, and the mRNA levels of ezh2 and core clock genes (clock1a, bmal1b, per1b, and per3) were measured at the ZT1 and ZT15 time points in 5 dpf larvae. The experiments were repeated three times. JTK-Cycle was used to analyse periodicity, amplitude, cycle, and phase. Bar scatter plots show cycle and amplitude data, and pie charts depict phase shifts; statistical significance was analysed for each time point (t-test: ns, p > 0.05; ∗, p < 0.05; ∗∗, p < 0.01; ∗∗∗, p < 0.001; ∗∗∗∗, p < 0.0001).