Mutagenesis Process and ENU

In the mouse, the supermutagen, N-ethyl-N-nitrosourea (ENU), produces forward mutation rates (∼1/650 per locus per gamete) sufficient to perform genome-wide mutagenesis screens at significant coverage so that, on average, mutations can be obtained in almost any gene. ENU mutagenesis screens provide a complimentary approach to gene targeting methods because both gain-of-function and loss-of-function alleles can be obtained, and importantly, no pre-existing knowledge of the target genes is required.

ENU causes single nucleotide mutations. The highest mutation rates occur in mouse pre-meiotic spermatogonial stem cells, with single-locus mutation frequencies of 6 x 10^-3 to 1.5 x 10^-3 per mutagenized genome (Hitotsumachi et al. 1985), depending on drug dose used (Justice 2000). Chemical mutagenesis by ENU has great potential to generate an abundance of mutations in phenotype-driven screens, given that each of the approximately 30,000 mammalian genes is a potential target for inducing a mutation with a demonstrable, heritable phenotype.

Large-scale ENU mutagenesis allows researchers access to more genetic variants from which to select mutants for further study, e.g., those with more striking phenotypes that are not inherently polygenic. Furthermore, the sequence of candidate genes or genomic sequence in critical regions between the mutated and parental strains can be more easily compared if the mutations are isolated on isogenic background. For these reasons, it has been suggested that gene discovery by ENU mutagenesis will be more successful than QTL approaches (e.g., Nadeau and Frankel 2000).