Disruptive selection, definition

Plasmids, such as RNAi transgenes, can be introduced into ES cells by several methods, including electroporation (42). A drug-selectable marker is convenient to have on the same plasmid as the U6 or HI promoter-driven shRNA. This allows for selection of stable, clonal ES cell lines that may be assayed for levels of knockdown and the generation of embryos and mice. An important consideration for RNAi transgenesis in ES cells is the dominant nature of the teclmique. The disruption of genes required for ES cell self-renewal or smvival, by definition, does not produce ES cell clones with significant knockdown. The ES cells will either differentiate or die, but if clones are recovered, they likely have mild to moderate levels of knockdown. In these situations, an inducible RNAi system is required to circumvent this problem, as will be discussed next. 

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