Details

Autor(en) / Beteiligte

• Wang, Jianbin
• Rebar, Edward J
• Guschin, Dmitry Y
• Lee, Ya-Li
• Holmes, Michael C
• Miller, Jeffrey C
• Gregory, Philip D
• Kim, Kenneth A
• Waite, Adam J
• Beausejour, Christian M
• Wang, Nathaniel S
• Pabo, Carl O
• Rupniewski, Igor

Titel

An improved zinc-finger nuclease architecture for highly specific genome editing

Ist Teil von

• Nature biotechnology, 2007-07, Vol.25 (7), p.778-785

Ort / Verlag

New York, NY: Nature Publishing Group

Erscheinungsjahr

2007

Link zum Volltext

Quelle

MEDLINE

Beschreibungen/Notizen

• Genome editing driven by zinc-finger nucleases (ZFNs) yields high gene-modification efficiencies (>10%) by introducing a recombinogenic double-strand break into the targeted gene. The cleavage event is induced using two custom-designed ZFNs that heterodimerize upon binding DNA to form a catalytically active nuclease complex. Using the current ZFN architecture, however, cleavage-competent homodimers may also form that can limit safety or efficacy via off-target cleavage. Here we develop an improved ZFN architecture that eliminates this problem. Using structure-based design, we engineer two variant ZFNs that efficiently cleave DNA only when paired as a heterodimer. These ZFNs modify a native endogenous locus as efficiently as the parental architecture, but with a >40-fold reduction in homodimer function and much lower levels of genome-wide cleavage. This architecture provides a general means for improving the specificity of ZFNs as gene modification reagents.