Supplementary Figure 4: Ectopic expression of abcc4 mRNA rescues lkt mutant phenotypes.

(a–d) lkt mutants displaying a ventrally curved axis (a), three otoliths (b; red arrows), hydrocephalus (c; red arrow) and reversed (left)-looped heart (d). (e–h) abcc4 mRNA injection rescued mutant phenotypes, including a straight body axis (e), two otoliths (f), normal brain (g) and normal (right)-looped heart (h). Scale bar: 100 μm (a–h). (i) Percentages of rescued lkt mutants grouped in heart looping, otolith biogenesis, axis formation or brain development. Approximately 160 pg of abcc4 mRNA was injected into embryos derived from crosses of lkt heterozygouts. Injected embryos were genotyped using SSLP markers (Z17212 and Z6907). Embryos linking with the lkt mutant genotype but showing wild-type phenotypes were scored as rescued mutants. %: The number of rescued lkt mutants divided by number of all lkt mutants. The total numbers of embryos pooled from n = 3 independent experiments are indicated. Graph shows mean ± s.d.; Student’s t test: ^∗∗∗P < 0.001. (j) Bar graph depicting percentages of normal (rescued) brain phenotypes in embryos co-injected with control morpholinos, antisense morpholinos and antisense morpholinos with specific mRNAs. Wild-type embryos were co-injected with cox1-MO (8 ng) with cox1 mRNA (130 pg), cox2-MO (8 ng) with cox2 mRNA (125 pg), ep4-MO (1 ng) with ep4 mRNA (120 pg) or abcc4-MO (12 ng) with abcc4 mRNA (160 ng). Ctrl: Ctrl-MO (8 ng). The total numbers of embryos pooled from n = 3 independent experiments are indicated. Graph shows mean ± s.d.; Student’s t test: ^∗∗∗P < 0.001. Statistics source data for Supplementary Fig. 4i, j can be found in Supplementary Table 2.