Journal
MOLECULAR CELL
Volume 31, Issue 2, Pages 294-301Publisher
CELL PRESS
DOI: 10.1016/j.molcel.2008.06.016
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Funding
- NCI NIH HHS [R01CA112021, R01 CA112021] Funding Source: Medline
- NCRR NIH HHS [R21 RR024189-01, R21RR024189, R21 RR024189] Funding Source: Medline
- NHLBI NIH HHS [R01 HL079295] Funding Source: Medline
- NIGMS NIH HHS [R24 GM078369-01A1, R24GM078369, R24 GM078369-02, R01 GM069906, R24 GM078369, R01 GM069906-05, R01 GM069906-04, R01GM069906, R01 GM069906-03] Funding Source: Medline
- NIH HHS [DP1 OD006862] Funding Source: Medline
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Custom-made zinc-finger nucleases (ZFNs) can induce targeted genome modifications with high efficiency in cell types including Drosophila, C. elegans, plants, and humans. A bottleneck in the application of ZFN technology has been the generation of highly specific engineered zinc-finger arrays. Here we describe OPEN (Oligomerized Pool ENgineering), a rapid, publicly available strategy for constructing multifinger arrays, which we show is more effective than the previously published modular assembly method. We used OPEN to construct 37 highly active ZFN pairs which induced targeted alterations with high efficiencies (1%-50%) at 11 different target sites located within three endogenous human genes (VEGF-A, HoxB13, and CFTR), an endogenous plant gene (tobacco SuRA), and a chromosomally integrated EGFP reporter gene. In summary, OPEN provides an open-source method for rapidly engineering highly active zinc-finger arrays, thereby enabling broader practice, development, and application of ZFN technology for biological research and gene therapy.
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