Targeted gene knockout by direct delivery of zinc-finger nuclease proteins

Abstract

Due to an unexpected cell-penetrating property, zinc-finger nucleases (ZFNs) can be delivered to several mammalian cell types as proteins. Dose-dependent disruption of an endogenous gene was achieved with reduced activity at known off-target sites. Zinc-finger nucleases (ZFNs) are versatile reagents that have redefined genome engineering. Realizing the full potential of this technology requires the development of safe and effective methods for delivering ZFNs into cells. We demonstrate the intrinsic cell-penetrating capabilities of the standard ZFN architecture and show that direct delivery of ZFNs as proteins leads to efficient endogenous gene disruption in various mammalian cell types with minimal off-target effects.