Blastocyst Microinjection with Embryonic Stem Cells

The ability to delete the function of an endogenous gene in the mouse was made possible by the development of embryonic stem (ES) cells, pluripotent cells that retain the ability to develop into all tissues of a developing embryo. The ability to genetically modify these cells followed, allowing targeted mutation of ES cells in vitro and the deletion of specific gene function. However, regardless of the simplicity or complexity of the genetic modification, all ES cells require injection into host embryos to establish pregnancies and result in chimeric mice. Blastocysts are commonly used as the host embryos for this purpose, as it is relatively easy to inject cells into the blastocoel cavity of the developing embryo. This chapter describes the procedure for injection of ES cells into blastocyst stage embryos for the generation of knockout mice.