Production of Mice using Chemical Mutagenesis of Mouse ES Cells
Mouse embryonic stem (ES) cells can contribute to all tissues during development, including the germline, permitting genetic modifications to be introduced in culture and analyzed in the mouse. Using mouse ES cells to combine advantages inherent to cell culture systems with chemical mutagenesis greatly enhances genetic approaches to address gene function. The ability to modulate and readily assay mutation rate, improved quality control, higher mutation frequency, a wider spectrum of mutagens and shortened breeding strategies in mice are attractive features of ES cell mutagenesis.
Mouse ES cells can be effectively mutagenized with the point mutagens ENU and ethylmethanesulfonate (EMS). ES cell mutagenesis permits a genome-wide recessive screen to be performed in two rather than three generations (Fig. 2). In the scheme shown, chimeric male mice derived from injecting blastocysts with chemically mutagenized ES cells are used to initiate the screen. The chimeric males are mated to wild-type females, and the G1 females are backcrossed to the original chimera. Alternatively, the G1 mice can be intercrossed to generate offspring that potentially carry recessive mutations (scheme not shown). In the breeding protocol shown 1/4 of the G2 mice will be homozygous for a mutation at a given locus, increasing the opportunity to identify a mutant as the G2 mice are channeled through the series of tests to detect a phenotype.
