Improving the sensitivity of ENU mutagenesis screens using inbred mice: implications for modifier gene discovery.

Positional cloning of ENU-induced mutations has traditionally relied on analysis of polymorphic variation between two strains. In contrast, the application of whole genome sequencing (WGS) has enabled gene discovery in mutant lines maintained on an inbred genetic background. This approach utilizes genetic variation derived from ENU-induced variants for mapping, and reduces the likelihood of phenotypic variation, making it an ideal method for genetic modifier screening. Here, we describe the results of such a screen, wherein we determined the minimal number of mutant genomic DNA samples to include in our analyses, and improved the sensitivity of our screen by individually barcoding each genomic DNA library. We present several unique cases to illustrate this approach’s efficacy, including the discovery of two distinct mutations that generate essentially identical mutant phenotypes, an approach to the identification of putative dominant genetic modifiers, and the ascertainment of a non-ENU-induced causal variant through homozygosity mapping. These results have increased our confidence that inbred screening followed by WGS analysis can be utilized to identify modifying loci.