title: A simple Gateway-assisted construction system of TALEN genes for plant genome editing.
abstract: TALEN is an artificial nuclease being applied for sequence-specific genome editing. For the plant genome editing, a pair of TALEN genes is expressed in the cells, and a binary plasmid for Agrobacterium-mediated transformation should be assembled. We developed a novel procedure using the Gateway-assisted plasmids, named Emerald-Gateway TALEN system. We constructed entry vectors, pPlat plasmids, for construction of a desired TALEN gene using Platinum Gate TALEN kit. We also created destination plasmid, pDual35SGw1301, which allowed two TALEN genes to both DNA strands to recruit using Gateway technology. Resultant TALEN genes were evaluated by the single-strand annealing (SSA) assay in E. coli cells. By this assay, the TALENs recognized the corresponding targets in the divided luciferase gene, and induced a specific recombination to generate an active luciferase gene. Using the TALEN genes constructed, we created a transformant potato cells in which a site-specific mutation occurred at the target site of the GBSS gene. This suggested that our system worked effectively and was applicable as a convenient tool for the plant genome editing.
authors.affiliation: Department of Biological Science and Technology, Tokyo University of Science, Katsushika, Tokyo 125-8585, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Kusano
authors.affiliation: Department of Biological Science and Technology, Tokyo University of Science, Katsushika, Tokyo 125-8585, Japan.
authors.forename: Hitomi
authors.initials: H
authors.lastname: Onodera
authors.affiliation: Department of Biological Science and Technology, Tokyo University of Science, Katsushika, Tokyo 125-8585, Japan.
authors.forename: Miho
authors.initials: M
authors.lastname: Kihira
authors.affiliation: Department of Biological Science and Technology, Tokyo University of Science, Katsushika, Tokyo 125-8585, Japan.
authors.forename: Hiromi
authors.initials: H
authors.lastname: Aoki
authors.affiliation: Department of Biological Science and Technology, Tokyo University of Science, Katsushika, Tokyo 125-8585, Japan.
authors.forename: Hikaru
authors.initials: H
authors.lastname: Matsuzaki
authors.affiliation: Department of Biological Science and Technology, Tokyo University of Science, Katsushika, Tokyo 125-8585, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Shimada
mesh_terms: D060054:Agrobacterium / Q000235:genetics; D004720:Endonucleases / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D000072669:Gene Editing; D018506:Gene Expression Regulation, Plant; D005822:Genetic Vectors; D018745:Genome, Plant / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011198:Solanum tuberosum / Q000235:genetics; D013214:Starch Synthase / Q000235:genetics*; D000069896:Transcription Activator-Like Effector Nucleases / Q000235:genetics*
chemical_list: D013214:Starch Synthase; D004720:Endonucleases; D000069896:Transcription Activator-Like Effector Nucleases