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稻曲病菌T-DNA插入突变体5062的插入位点分析
引用本文:俞咪娜,胡建坤,黄磊,于俊杰,尹小乐,聂亚锋,陈志谊,刘永锋. 稻曲病菌T-DNA插入突变体5062的插入位点分析[J]. 中国农业科学, 2013, 46(9): 1790-1798. DOI: 10.3864/j.issn.0578-1752.2013.09.006
作者姓名:俞咪娜  胡建坤  黄磊  于俊杰  尹小乐  聂亚锋  陈志谊  刘永锋
作者单位:江苏省农业科学院植物保护研究所,南京 210014
基金项目:国家自然科学基金(31171802)、江苏省农业自主创新基金(CX(12)1003-10)
摘    要:【目的】研究稻曲病菌致病力增强的ATMT突变菌株5062,为稻曲病菌致病机制解析、致病相关基因克隆提供方法基础。【方法】测定和观察5062的菌落直径、菌落形态、产孢能力等生物学特性,进一步利用TAIL-PCR和RACE技术克隆5062基因组的T-DNA侧翼基因,并比对分析基因的信息,最后利用RT-PCR检测突变基因的表达量。【结果】与野生型菌株相比,突变菌株5062田间接种表现为致病力增强,在MM和水琼脂培养基上生长速率减慢,产生大量分生孢子,而在PSA和TB3培养基上,其菌落形态、色素等方面与野生型无明显差异。TAIL-PCR扩增得到的紧邻T-DNA两侧的侧翼序列在野生型中不相邻。TAIL-PCR结合RACE技术克隆了5062基因组的T-DNA侧翼基因,RT-PCR表明T-DNA插入位点右侧1个基因在突变体中上调表达。【结论】突变菌株5062的致病力增强,在营养贫乏条件下产孢能力增强,其表型的变化可能是染色体重排和T-DNA插入共同影响的结果。

关 键 词:稻曲病菌   致病力   T-DNA   突变   染色体重排
收稿时间:2012-12-24

Molecular Characterization of T-DNA Integration of the Ustilaginoidea virens Mutant 5062
YU Mi-Na,HU Jian-Kun,HUANG Lei,YU Jun-Jie,YIN Xiao-Le,NIE Ya-Feng,CHEN Zhi-Yi,LIU Yong-Feng. Molecular Characterization of T-DNA Integration of the Ustilaginoidea virens Mutant 5062[J]. Scientia Agricultura Sinica, 2013, 46(9): 1790-1798. DOI: 10.3864/j.issn.0578-1752.2013.09.006
Authors:YU Mi-Na  HU Jian-Kun  HUANG Lei  YU Jun-Jie  YIN Xiao-Le  NIE Ya-Feng  CHEN Zhi-Yi  LIU Yong-Feng
Affiliation:Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing 210014
Abstract:【Objective】 The objective of the experiment is to establish a method for cloning genes of the Ustilaginoidea virens mutant, and to shed light on the pathogenic mechanism of the U. virens, depending on a new mutant 5062, which has increased virulence. 【Method】With the wild type strain 70-22 as a control, phenotypic analysis, such as growth rate, colonial morphologies and sporulation, were analyzed. TAIL-PCR combined with RACE were used to identify the T-DNA integration site and the genes of flanking right site of the T-DNA. The expression levels of the genes were analyzed by RT-PCR as well. 【Result】Colonial morphologies and pigment on PSA and TB3 medium were no different between 70-22 and 5062. But on MM and water-agar medium, 5062 grew slower and produced more conidiophores. The flanking sequences of T-DNA were non-adjacent in the wild type. The expression of the gene flanking right site of the T-DNA was upregulated.【Conclusion】In 5062, both the T-DNA insertion and inversion of chromosomal fragment caused a mutation of the increased virulence.
Keywords:Ustilaginoidea virens  virulence  T-DNA  mutant  chromosomal rearrangement
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