title: Targeting DNA to a previously integrated transgenic locus using zinc finger nucleases.
abstract: Targeting exogenously supplied DNA to a predetermined location within a plant genome provides a powerful tool for basic studies of plant gene function and opens up some intriguing possibilities for crop improvement. The induction of double-strand DNA breaks at specific genomic loci via the use of designed zinc finger nucleases (ZFNs) allows for targeted transgene integration. Preintegrating a reporter construct containing a nonfunctional herbicide resistance gene flanked by ZFN binding sites results in a locus capable of being targeted. Retransformation with a corresponding ZFN-expressing cassette and a donor DNA with sequences homologous to the integrated construct and capable of functionalizing the herbicide resistance gene following site-specific integration results in targeted DNA addition. Targeted DNA integration can be confirmed in herbicide-resistant plant cells using PCR analysis.
authors.affiliation: Discovery R&D, Dow AgroSciences, Indianapolis, IN, USA.
authors.forename: Tonya L
authors.initials: TL
authors.lastname: Strange
authors.forename: Joseph F
authors.initials: JF
authors.lastname: Petolino
mesh_terms: D000123:Acetyltransferases / Q000235:genetics*; D053903:DNA Breaks, Double-Stranded; D018744:DNA, Plant / Q000235:genetics; D004720:Endonucleases / Q000235:genetics / Q000378:metabolism*; D018390:Gene Targeting / Q000379:methods*; D018014:Gene Transfer Techniques; D017343:Genes, Plant; D018745:Genome, Plant; D053209:Herbicide Resistance / Q000235:genetics*; D006540:Herbicides / Q000494:pharmacology; D016254:Mutagenesis, Insertional; D009943:Organophosphorus Compounds / Q000494:pharmacology; D030821:Plants, Genetically Modified / Q000235:genetics; D046509:Tissue Culture Techniques; D014026:Nicotiana / Q000235:genetics*; D019076:Transgenes; D016335:Zinc Fingers / Q000235:genetics*
chemical_list: D018744:DNA, Plant; D006540:Herbicides; D009943:Organophosphorus Compounds; C003122:bialaphos; D000123:Acetyltransferases; C056243:phosphinothricin N-acetyltransferase; D004720:Endonucleases