Extended Data Fig. 6: Knockdown of upf1, upf2 or upf3b has no effects on GCR in both capn3a^−/− and nid1a^−/− embryos.

a–f, WISH was performed to assess the liver sizes in 3.5-dpf wild-type and capn3a^−/− embryos injected with different reagents as indicated (a, c, e). Statistical analysis of the average liver sizes corresponding to a, c and e is shown in b, d and f, respectively. g, Relative mRNA expression of capn3a, capn8 and capn12 in wild-type and capn3a^−/− embryos injected with or without upf2-1#MO at 1.5 dpf. h, Left, validation of the second gene-specific morpholino of upf2 (upf2-2#MO). The upf2-2#MO was designed to block the splicing site of exon3–intron3 of upf2. RT–PCR was performed with a pair of primers (pf, forward primer; pv, reverse primer) to distinguish the mRNA produced by the wild-type embryos injected with ST-MO from that injected with upf2-2#MO. DNA fragment of 1,638 bp contains 105 bp of exon 3, 61 bp of exon 4 and the whole intron 3 of upf2. Right, relative mRNA expression of capn3a, capn8 and capn12 in wild-type and capn3a^−/− embryos injected with different morpholinos as indicated at 1.5 dpf. i, j, Relative mRNA expression of capn3a, capn8 and capn12 in wild-type embryos, wild-type embryos injected with upf3b-MO (i) or upf1-MO (j), capn3a^−/− embryos, or capn3a^−/− embryos injected with upf3b-MO or upf1-MO at 1.5 dpf. k–m, Statistical analysis of the average body lengths in 4-dpf wild-type and nid1a^−/− larvae injected with or without different morpholinos as indicated. n, Relative mRNA expression of endogenous capn3a in Tg1-1# and Tg3-1# embryos injected with or without upf3a-MO. Each experiment was repeated three times with similar results, and a representative result is shown. n indicates the number of zebrafish embryos in each group. Data are mean ± s.d. from three biological replicates. *P < 0.05, **P < 0.01, ***P < 0.001, two-tailed t-test.