Fig. 1. Expression of Barhl1 in the developing inner ear and targeted disruption of the Barhl1 locus. (A-C) RNA in situ hybridization analysis of coronal ear sections shows signals in the utricular maculae, cristae of semicircular canals and cochlear hair cells (arrows). (D,E) ß-galactosidase staining (counterstained with Fast Red) of Barhl1^+/– inner ear sections labels cochlear outer and inner hair cells, and saccular hair cells. (F,G) ß-galactosidase staining of Barhl1^+/– whole-mount organs of Corti and cristae at P5: note the much more intense signals in cochlear outer hair cells compared with inner hair cells and hair cells of the crista. (H) Schematic of the inner ear, indicating the expression levels of Barhl1 in the cochlea and vestibule. (I) Homologous recombination between the wild-type allele and the targeting vector results in the replacement of Barhl1-coding exons (black boxes) with the lacZ marker DNA and a PGK-Neo cassette flanked by two loxp sites (hatched boxes). E, EcoRI; H, HindIII; K, KpnI; S, SalI; ScI, SacI; ScII, SacII; Xb, XbaI; Xh, XhoI. (J) Southern blot analysis of SacI-digested DNA from wild-type, heterozygous and homozygous mutant mice. The 3' probe identifies SacI fragments of 7.8 kb (wild-type allele) and 4.5 kb (targeted allele). (K) PCR analysis of DNA from wild-type, heterozygous and homozygous mutant mice. The wild-type and targeted alleles yield a product of 350 bp and 730 bp (derived from lacZ), respectively. AVC, anterior vertical canal; Co, cochlea; Cr, crista; HC, horizontal canal; IHC, inner hair cell; OHC, outer hair cell; PVC, posterior vertical canal; S, saccule; U, utricule. Scale bars: 50 µm.

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