Molecular characterisation of the Hyp deletion and an improved assay for its detection

The Hyp mouse is a commonly used model for the study of the phosphate wasting disease X-linked hypophosphataemia. The defect in this mouse line is a deletion that includes exons 16 to 22 of Phex, although the exact extent of this X chromosome deletion remains unknown. This complicates genotyping which increases costs, time and difficulty of working with this important model. We aimed to determine the molecular breakpoints of this deletion in order develop a robust assay for its detection. We designed short mapping PCRs around the Phex locus to refine the putative breakpoint locations, then used gap PCR to amplify a product containing the breakpoint junction. DNA sequencing showed the deleted region was approximately 297 kb, significantly larger than previous reports, but did not contain any genes other than Phex. DNA sequence analysis revealed that this deletion may be the result of microhomology-mediated end joining. Finally, we designed a multiplex PCR assay for genotyping Hyp colonies and validated it using a panel of Hyp colony mice. This study provides confirmation of the Hyp phenotype as a single gene defect, a potential mechanism for its formation and an improved method for genotyping that will make working with this strain significantly easier.

► The deletion in the Hyp mouse model of X-linked hypophosphataemia was characterised.
► The Hyp deletion was more extensive than previously described but only affects Phex.
► A genotyping method for direct detection of the Hyp deletion was established.