Production of Mice for a Genome-Wide ENU Mutagenesis Screen
ENU induced mutagenesis is performed using predominantly C57BL/6J and a smaller percentage of A.B6-Tyr+ mice. These injected males represent G0 mice; their male offspring represents G1 (founder) mice, which represent the equivalent of one mutagenized gamete (genome) captured in the mating of a wild-type female to an ENU-treated G0 male. Since the NMF is most interested in identifying genome-wide recessive mutations, the production and screening of third-generation (G3) offspring from the G1 founder, is necessary. Thus, in a genome-wide, recessive screen the two critical classes are the G1 mice (= # of mutagenized genomes screened), and the G3 mice (= # of animals channeled through phenotyping). In the backcross breeding scheme (Fig.1), all of the G3 progeny will be recessive for a subset of the mutations induced in the G1 founder with any specific mutation represented in 1/8 G3 offspring. (Dominant mutations are also detected in the G3). The planned production of 20 G3 progeny for each G1 provides a 92% chance that a new mutation will be seen at least once and a 71% chance that it will be seen two or more times - providing valuable insight on heritability.
We will produce and screen at least 8,000 G3 progeny per year. At 20:1 G3/G1 this reflects an annual total of 440 mutagenized genomes. Assuming 100,000 genes and an average locus mutation rate of between 1/500-1,000 this represents 1x access to ~40-80% of recessive mutations and ~8-16% of dominants. The backcross breeding strategy outlines the production of 200 G3 mice each week to provide a steady supply of mice for phenotyping.
