Preparation of DNA from Embryonic Stem Cells or Other Cultured Cells

Abstract

Characterization of embryonic stem (ES) cell-mediated genome alterations, including random insertional transgenesis, gene trapping, gene targeting, and site-specific recombinase-mediated changes, is performed mostly at the ES cell level, before the introduction of these alterations into a mouse. A detailed characterization requires a larger amount of DNA than is required for the initial detection of the candidates for the desired alteration. This protocol describes the preparation of DNA from a 10-cm tissue culture plate. The cells are trypsinized and lysed, and DNA is recovered from the lysate by organic extraction followed by ethanol precipitation.