title: A simple method for construction of artificial microRNA vector in plant.
abstract: Artificial microRNA (amiRNA) is a powerful tool for silencing genes in many plant species. Here we provide an easy method to construct amiRNA vectors that reinvents the Golden Gate cloning approach and features a novel system called top speed amiRNA construction (TAC). This speedy approach accomplishes one restriction-ligation step in only 5 min, allowing easy and high-throughput vector construction. Three primers were annealed to be a specific adaptor, then digested and ligated on our novel vector pTAC. Importantly, this method allows the recombined amiRNA constructs to maintain the precursor of osa-miR528 with exception of the desired amiRNA/amiRNA* sequences. Using this method, our results showed the expected decrease of targeted genes in Nicotiana benthamiana and Oryza sativa.
authors.affiliation: State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, Wuhan, 430072, China, wwwhpuedu@whu.edu.cn.
authors.forename: Yang
authors.initials: Y
authors.lastname: Li
authors.forename: Yang
authors.initials: Y
authors.lastname: Li
authors.forename: Sunping
authors.initials: S
authors.lastname: Zhao
authors.forename: Sheng
authors.initials: S
authors.lastname: Zhong
authors.forename: Zhaohai
authors.initials: Z
authors.lastname: Wang
authors.forename: Bo
authors.initials: B
authors.lastname: Ding
authors.forename: Yangsheng
authors.initials: Y
authors.lastname: Li
mesh_terms: D003001:Cloning, Molecular; D018744:DNA, Plant / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics; D029721:Drosophila Proteins / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D005822:Genetic Vectors; D035683:MicroRNAs / Q000235:genetics*; D012275:Oryza / Q000235:genetics*; D014026:Nicotiana / Q000235:genetics*
chemical_list: D018744:DNA, Plant; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D035683:MicroRNAs; C117995:osa protein, Drosophila