title: An improved particle bombardment for the generation of transgenic plants by direct immobilization of relleasable Tn5 transposases onto gold particles.
abstract: We have developed a modified particle bombardment method for plant transgenesis. An intein-tag and a 6×Cys-tag were successively fused to the N-terminus of a hyperactive Tn5 transposase. The modified transposase was immobilized on bare gold microscopic particles via covalent binding of a 6×Cys-tag sulfydryl groups to the gold surface. The tethered transposase can bind the transposon DNA in vitro to form the transposome in the absence of Mg²⁺ ions. After bombardment of the gold particles carrying the transposomes into the plant cells, the transposomes will be released from the carrier due to the activated self-cleavage function of intein-tag. Our data showed this procedure integrated foreign DNA into the plant genome with an increased transformation frequency as compared to the conventional particle bombardment method. A single copy insertion can also be obtained by decreasing of the assembled transposon DNA amount in relation to plant cell biomass.
authors.affiliation: State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing 210046, People's Republic of China.
authors.forename: Jun
authors.initials: J
authors.lastname: Wu
authors.forename: Hongwei
authors.initials: H
authors.lastname: Du
authors.forename: Xuewei
authors.initials: X
authors.lastname: Liao
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhao
authors.forename: Liguan
authors.initials: L
authors.lastname: Li
authors.forename: Liuyan
authors.initials: L
authors.lastname: Yang
mesh_terms: D005818:Genetic Engineering / Q000379:methods*; D006046:Gold / Q000737:chemistry*; D008957:Models, Genetic; D029595:Orchidaceae / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics*; D014170:Transformation, Genetic; D019895:Transposases / Q000737:chemistry / Q000235:genetics*; D003313:Zea mays / Q000235:genetics
chemical_list: C473503:Tn5 transposase; D006046:Gold; D019895:Transposases