Targeted molecular trait stacking in cotton through targeted double‐strand break induction

作者：Kathleen D’Halluin, Chantal Vanderstraeten, Jolien Van Hulle, Joanna Rosolowska, Ilse Van den Brande, Anouk Pennewaert, Kristel D'Hont, Martine Bossut, Derek Jantz, René K. Ruiter, Jean Broadhvest · 发表于：Plant Biotechnology Journal · 年份：2013 · DOI：10.1111/pbi.12085 · 被引用次数：162 · 研究领域：CRISPR and Genetic Engineering、Plant Virus Research Studies、Plant tissue culture and regeneration

Recent developments of tools for targeted genome modification have led to new concepts in how multiple traits can be combined. Targeted genome modification is based on the use of nucleases with tailor-made specificities to introduce a DNA double-strand break (DSB) at specific target loci. A re-engineered meganuclease was designed for specific cleavage of an endogenous target sequence adjacent to a transgenic insect control locus in cotton. The combination of targeted DNA cleavage and homologous recombination-mediated repair made precise targeted insertion of additional trait genes (hppd, epsps) feasible in cotton. Targeted insertion events were recovered at a frequency of about 2% of the independently transformed embryogenic callus lines. We further demonstrated that all trait genes were inherited as a single genetic unit, which will simplify future multiple-trait introgression.