The derivation of embryonic stem (ES) cells represents one of the most important breakthroughs in mammalian developmental biology. In addition to their utility in a wide array of in vitro studies, ES cells are also one of the most useful starting materials for the generation of mutants by homologous recombination in mice (Thomson and Solter, 1988). When ES cells are injected into host blastocysts and transferred to the uterus of a pseudo-pregnant mouse, they can contribute to different types of tissues in chimeric mice, including the germ line (Bradley et al., 1984). Hundreds of genes have been studied through genetic manipulation of ES cells to model human genetic diseases. In this unit, the ES cell lines are derived from the 129SvEv mice strain, which has a high probability of promoting germ line transmission. Procedures for validating and characterizing ES cell pluripotency are also described in detail.
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