A Simple Method that Combines CRISPR and AID to Quickly Generate Conditional Knockouts for Essential Genes in Various Vertebrate Cell Lines

Abstract

Cells with a loss-of-function mutation in a gene (knockout cells) are powerful tools for characterizing the function of such gene product. However, for essential genes, conditional knockout cell lines must be generated. The auxin-inducible degron (AID) technique enables us to conditionally and rapidly deplete a target protein from various eukaryotic cell lines. A combination of CRISPR-/Cas9-based gene editing and AID technique allows us to generate AID-based conditional knockout cell lines. Using these two techniques, we recently proposed a simple and quick way to generate conditional knockout cells for essential genes. In this chapter, we introduce the reader to the experimental procedures to generate these AID-based conditional knockout cell lines.

Keywords: Auxin-Inducible Degron (AID); CRISPR/Cas9; Conditional knockout; Essential genes.