Agrobacterium tumefaciens-mediated transformation: An efficient tool for insertional mutagenesis and targeted gene disruption in Harpophora oryzae

文献类型： 外文期刊

第一作者： Liu, Ning

作者： Liu, Ning;Ning, Guo-Ao;Shi, Huan-Bin;Zhang, Chu-Long;Feng, Xiao-Xiao;Liu, Xiao-Hong;Su, Zhen-Zhu;Lin, Fu-Cheng;Chen, Guo-Qing;Lu, Jian-Ping;Lin, Fu-Cheng;Mao, Li-Juan

作者机构：

关键词： Endophytic fungus;Harpophora oryzae;Agrobacterium tumefaciens-mediated;transformation;Insertional mutagenesis;Gene deletion mutagenesis;Sulfonylurea resistance

期刊名称：MICROBIOLOGICAL RESEARCH （ 影响因子：5.415； 五年影响因子：6.038 ）

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收录情况： SCI

摘要： The endophytic filamentous fungus Harpophora oryzae is a beneficial endosymbiont isolated from the wild rice. H. oryzae could not only effectively improve growth rate and biomass yield of rice crops, but also induce systemic resistance against the rice blast fungus, Magnaporthe oryzae. In this study, Agrobacterium tumefaciens-mediated transformation (ATMT) was employed and optimized to modify the H. oryzae genes by either random DNA fragment integration or targeted gene replacement. Our results showed that co-cultivation of H. oryzae conidia with A. tumefaciens in the presence of acetosyringone for 48 h at 22 degrees C could lead to a relatively highest frequency of transformation, and 200 acetosyringone (AS) pre-cultivation of A. tumefaciens is also suggested. ATMT-mediated knockout mutagenesis was accomplished with the gene-deletion cassettes using a yeast homologous recombination method with a yeast-Escherichia-Agrobacterium shuttle vector pKOHo. Using the ATMT-mediated knockout mutagenesis, we successfully deleted three genes of H. oryzae (HoATG5, HoATG7, and HoATG8), and then got the null mutants Delta Hoatg5, Delta Hoatg7, and Delta Hoatg8. These results suggest that ATMT is an efficient tool for gene modification including randomly insertional mutagenesis and gene deletion mutagenesis in H. oryzae. (c) 2015 Elsevier GmbH. All rights reserved.

分类号： Q

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