Mutation identification. (A) Exome sequencing. Graphical representation of sequence tags from exome sequencing (exon 5) of a kindred 1 HX patient (left), where 17 of 31 full-length reads have an A instead of a G, leading to a missense mutation (Val to Met at amino acid 282), and a kindred 2 HX patient (right), where 41 of 84 full-length reads have an A instead of a T, leading to a missense mutation (Val to Glu at amino acid 282). (B) Sanger sequencing confirmation of KCNN4 gene mutations showing a partial exon 5 wild-type sequence (top), the corresponding sequence from a kindred 1 HX heterozygote (middle), and the corresponding sequence from a kindred 2 HX heterozygote (bottom). (C) Conservation of mutations across vertebrate species. The mutant amino acid residue identified in both HX patients is conserved across vertebrate species, including the clades of placental mammals, the extant Eutherians, and within members of the small-intermediate family of proteins.