Homologous recombination: a basis for targeted genome optimization in crop species such as maize.
In an attempt to understand the feasibility of future targeted genome optimization in agronomic crops, we tested the efficiency of homologous recombination-mediated sequence insertion upon induction of a targeted DNA double-strand break at the desired integration site in maize. By the development of an efficient tissue culture protocol, and with the use of an I-SceI gene optimized for expression in maize, large numbers of precisely engineered maize events were produced in which DNA integration occurred very accurately. In a subset of events examined in detail, no additional deletions and/or insertions of short filler DNA at the integration site were observed. In 30%-40% of the recovered events, no traces of random insertions were observed. This was true for DNA delivery by both Agrobacterium and particle bombardment. These data suggest that targeted double-strand break-induced homologous recombination is a superior method to generate specific desired changes in the maize genome, and suggest targeted genome optimization of agronomic crops to be feasible.
D'Halluin
Kathleen
K
Bayer BioScience N.V., Technologiepark 38, B-9052 Gent, Belgium. kathleen.d'halluin@bayercropscience.com
Vanderstraeten
Chantal
C
Stals
Ellen
E
Cornelissen
Marc
M
Ruiter
Rene
R
D000613:Aminobutyrates; D053903:DNA Breaks, Double-Stranded; D015252:Deoxyribonucleases, Type II Site-Specific / Q000235:genetics*; D018390:Gene Targeting / Q000379:methods*; D018745:Genome, Plant; D053209:Herbicide Resistance; D006540:Herbicides; D011995:Recombination, Genetic; D029701:Saccharomyces cerevisiae Proteins; D003313:Zea mays / Q000235:genetics*
D000613:Aminobutyrates; D006540:Herbicides; D029701:Saccharomyces cerevisiae Proteins; C003121:phosphinothricin; C037725:SCEI protein, S cerevisiae; D015252:Deoxyribonucleases, Type II Site-Specific