title: A Novel Phenolic Compound, Chloroxynil, Improves Agrobacterium-Mediated Transient Transformation in Lotus japonicus.
abstract: Agrobacterium-mediated transformation is a commonly used method for plant genetic engineering. However, the limitations of Agrobacterium host-plant interactions and the complexity of plant tissue culture often make the production of transgenic plants difficult. Transformation efficiency in many legume species, including soybean and the common bean, has been reported to be quite low. To improve the transformation procedure in legumes, we screened for chemicals that increase the transformation efficiency of Lotus japonicus, a model legume species. A Chemical library was screened and chemicals that increase in transient transformation efficiency of L. japonicus accession, Miyakojima MG-20 were identified. The transient transformation efficiency was quantified by reporter activity in which an intron-containing reporter gene produces the GUS protein only when the T-DNA is expressed in the plant nuclei. We identified a phenolic compound, chloroxynil, which increased the genetic transformation of L. japonicus by Agrobacterium tumefaciens strain EHA105. Characterization of the mode of chloroxynil action indicated that it enhanced Agrobacterium-mediated transformation through the activation of the Agrobacterium vir gene expression, similar to acetosyringone, a phenolic compound known to improve Agrobacterium-mediated transformation efficiency. Transient transformation efficiency of L. japonicus with 5 μM chloroxynil was 60- and 6- fold higher than that of the control and acetosyringone treatment, respectively. In addition, transgenic L. japonicus lines were successfully generated by 5 μM chloroxynil treatment.Furthermore, we show that chloroxynil improves L. japonicus transformation by Agrobacterium strain GV3101 and rice transformation. Our results demonstrate that chloroxynil significantly improves Agrobacterium tumefaciens-mediated transformation efficiency of various agriculturally important crops.
authors.affiliation: Department of Frontier Research, Kazusa DNA Research Institute, Kisarazu, Chiba, Japan.
authors.forename: Mitsuhiro
authors.initials: M
authors.lastname: Kimura
authors.affiliation: Department of Botany and Plant Sciences, Center for Plant Cell Biology and Institute for Integrative Genome Biology, University of California Riverside, Riverside, California, United States of America.
authors.forename: Sean
authors.initials: S
authors.lastname: Cutler
authors.affiliation: Department of Frontier Research, Kazusa DNA Research Institute, Kisarazu, Chiba, Japan.
authors.forename: Sachiko
authors.initials: S
authors.lastname: Isobe
mesh_terms: D000098:Acetophenones / Q000737:chemistry / Q000494:pharmacology; D016960:Agrobacterium tumefaciens / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D006540:Herbicides / Q000633:toxicity; D000070116:Lotus / Q000187:drug effects / Q000235:genetics / Q000254:growth & development*; D009570:Nitriles / Q000737:chemistry / Q000494:pharmacology*; D012275:Oryza / Q000187:drug effects / Q000235:genetics / Q000254:growth & development; D018515:Plant Leaves / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics; D014170:Transformation, Genetic / Q000187:drug effects*
chemical_list: D000098:Acetophenones; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D006540:Herbicides; D009570:Nitriles; D010940:Plant Proteins; C036483:T-DNA; C051667:acetosyringone; C584256:chloroxynil