Analysis of compound heterozygotes reveals that the mouse floxed Pax6 tm1Ued allele produces abnormal eye phenotypes

Abstract

Analysis of abnormal phenotypes produced by different types of mutations has been crucial for our understanding of gene function. Some floxed alleles that retain a neomycin-resistance selection cassette (neo cassette) are not equivalent to wild-type alleles and provide useful experimental resources. Pax6 is an important developmental gene and the aim of this study was to determine whether the floxed Pax6tm1Ued (Pax6fl) allele, which has a retained neo cassette, produced any abnormal eye phenotypes that would imply that it differs from the wild-type allele. Homozygous Pax6fl/fl and heterozygous Pax6fl/+ mice had no overt qualitative eye abnormalities but morphometric analysis showed that Pax6fl/fl corneas tended be thicker and smaller in diameter. To aid identification of weak effects, we produced compound heterozygotes with the Pax6Sey-Neu (Pax6−) null allele. Pax6fl/− compound heterozygotes had more severe eye abnormalities than Pax6+/− heterozygotes, implying that Pax6fl differs from the wild-type Pax6+ allele. Immunohistochemistry showed that the Pax6fl/− corneal epithelium was positive for keratin 19 and negative for keratin 12, indicating that it was abnormally differentiated. This Pax6fl allele provides a useful addition to the existing Pax6 allelic series and this study demonstrates the utility of using compound heterozygotes with null alleles to unmask cryptic effects of floxed alleles.