Targeted transgene integration in plant cells using designed zinc finger nucleases

被引：157

作者：

Cai, Charles Q. ^{[1
]}

Doyon, Yannick ^{[2
]}

Ainley, W. Michael ^{[1
]}

Miller, Jeffrey C. ^{[2
]}

DeKelver, Russell C. ^{[2
]}

Moehle, Erica A. ^{[2
]}

Rock, Jeremy M. ^{[2
]}

Lee, Ya-Li ^{[2
]}

Garrison, Robbi ^{[1
]}

Schulenberg, Lisa ^{[1
]}

Blue, Ryan ^{[1
]}

Worden, Andrew ^{[1
]}

Baker, Lisa ^{[1
]}

Faraji, Farhoud ^{[2
]}

Zhang, Lei ^{[2
]}

Holmes, Michael C. ^{[2
]}

Rebar, Edward J. ^{[2
]}

Collingwood, Trevor N. ^{[2
]}

Rubin-Wilson, Beth ^{[1
]}

Gregory, Philip D. ^{[2
]}

Urnov, Fyodor D. ^{[2
]}

Petolino, Joseph F. ^{[1
]}

机构：

[1] Dow AgroSci LLC, Indianapolis, IN 46268 USA

[2] Sangamo BioSci, Richmond, CA 94804 USA

来源：

PLANT MOLECULAR BIOLOGY | 2009年 / 69卷 / 06期

关键词：

Double strand DNA breaks; Homology-directed repair; Site-directed transgene integration; Zinc finger nucleases; DOUBLE-STRAND BREAKS; AGROBACTERIUM-MEDIATED TRANSFORMATION; HOMOLOGOUS RECOMBINATION; ARABIDOPSIS-THALIANA; GENE; EXPRESSION; GENOME; RICE; REPAIR; DISRUPTION;

D O I：

10.1007/s11103-008-9449-7

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

Targeted transgene integration in plants remains a significant technical challenge for both basic and applied research. Here it is reported that designed zinc finger nucleases (ZFNs) can drive site-directed DNA integration into transgenic and native gene loci. A dimer of designed 4-finger ZFNs enabled intra-chromosomal reconstitution of a disabled gfp reporter gene and site-specific transgene integration into chromosomal reporter loci following co-transformation of tobacco cell cultures with a donor construct comprised of sequences necessary to complement a non-functional pat herbicide resistance gene. In addition, a yeast-based assay was used to identify ZFNs capable of cleaving a native endochitinase gene. Agrobacterium delivery of a Ti plasmid harboring both the ZFNs and a donor DNA construct comprising a pat herbicide resistance gene cassette flanked by short stretches of homology to the endochitinase locus yielded up to 10% targeted, homology-directed transgene integration precisely into the ZFN cleavage site. Given that ZFNs can be designed to recognize a wide range of target sequences, these data point toward a novel approach for targeted gene addition, replacement and trait stacking in plants.

引用

页码：699 / 709

页数：11

共 45 条

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