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Fig. S3 Rescue with a splice-blocking morpholino restores nkcc1 transcript in the ear. (A-E) Whole-mount in situ hybridisation at 6 dpf with an nkcc1 probe shows a graded reappearance of transcript in the ears of to27d embryos injected with 4 ng splice-blocking morpholino, which correlates with the ear size in the live embryo. In the morpholino-rescued mutant embryos, wild-type transcript levels (c.f. uninjected sib, A) are not seen; however, the mild (C), moderate (D) and severe (E) ear volume reduction classes have increased nkcc1 levels when compared with the uninjected mutant (B). All embryos shown were stained under identical conditions and the reaction was stopped simultaneously. (F-J) Confirmation of the genotype of the embryos shown in A-E by sequencing. A is a heterozygous sib (F); B-E are homozygous mutants. Asterisks denote critical nucleotide. h, hub of semicircular canal. Scale bar: 50 μm.