From phenotype to the causal mutation: recessive defects

Abstract

This article presents the methodology used to identify the mutation responsible for a genetic defect from the observation of cases. In the first step, a set of homogeneous cases are collected, from the same breed and with the same clinical signs. This collection is completed by a related but unaffected control population. A pedigree analysis is possible to point towards a common ancestor who may have transmitted the defect to all cases. A genotyping step using a SNP chip is used to display a small genomic region homozygous and identical at all markers and including the mutation. The mutation is then identified by genome sequencing of a few cases followed by the filtering of the variants against a large sequence database of unaffected animals. The best candidate variants are retained on the basis of their functional annotation. The mutation is then statistically confirmed on the basis of large scale genotyping, to verify the complete association between the mutation and the phenotype. Finally, the causality of the mutation is proven by functional analysis, including RNA and protein analysis, cellular imaging, and even through transgenic models carrying the mutation.