A. miRNA expression profiling in lin-28a and lin-28b morphants embryos at 5 hpf. Hierarchically clustered heat-map representing differences in miRNA expression between control MO-injected embryos and lin-28a MO- or lin-28b MO-injected embryos. Relative levels of expression are colored from green (low) to red (high). B. Real-time PCR analysis of let-7a, let-7b, miR-430a and miR-430b expression in control MO-, lin-28a MO- and lin-28b MO-injected embryos. Expression levels of let-7a, let-7b, miR-430a and miR-430b were analyzed at 5 hpf using TaqMan miRNA assay. The expression level of miR-430a and miR-430b was downregulated in both lin-28a MO- and lin-28b MO-injected embryos compared with control MO-injected embryos (***p<0.001, control MO vs. lin-28a MO, control MO vs. lin-28b MO, two-tailed test, n = 3, mean ± SEM). C. Real-time PCR analysis of let-7a and let-7b expression in control MO-, lin-28a MO- and lin-28b MO-injected embryos. Expression levels of let-7a and let-7b were analyzed using TaqMan miRNA assay at 28 hpf. The expression level of let-7a and let-7b was increased in both lin-28a MO- and lin-28b MO-injected embryos embryos compared with control MO-injected embryos (*p<0.05, **p<0.01, control MO vs. lin-28a MO, control MO vs. lin-28b MO, two-tailed test, n = 4, mean ± SEM). D. Semiquantitative RT-PCR analysis of lin-41 gene expression in control MO-, lin-28a MO- and lin-28b MO-injected embryos. Total RNA was isolated from lin-28a MO-, lin-28b MO- or control MO-injected embryos at 24 hpf and analyzed by using specific primers for lin-41. The expression of lin-41 was decreased in both lin-28a MO- and lin-28b MO-injected embryos compared with control MO-injected embryos. E. Control, lin-28a and lin-28b MO-injected embryos labeled by in situ hybridization with lin-41 probe. Expression of lin-41 was repressed in CNS of both lin-28a and lin-28b morphant embryos at 24 hpf.
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