苹果树腐烂病菌T-DNA插入突变体的筛选及Vmzfp3基因的功能分析

发布时间：2018-06-18 17:36

本文选题：苹果树腐烂病 + 锌指蛋白　；参考：《河北农业大学学报》2017年02期

【摘要】：本研究采用接种离体枝条的方法,对150个苹果树腐烂病菌的T-DNA插入突变体库中的突变体进行了致病性的筛选,经过多次试验得到了3个致病力明显减弱的突变体,编号为X4379,X4368,X4468。基因组重测序结果显示,突变体X4379有2个T-DNA插入位点,分别是锌指蛋白3(Vmzfp3)和跨膜蛋白42(Vmtme42)。以苹果树腐烂病菌的全基因组序列为依托,设计引物,构建敲除载体以及互补表达载体。运用PEG的方法转化苹果树腐烂病菌,分别获得Vmzfp3和Vmtme42的敲除转化子和互补转化子。对Vmzfp3和Vmtme42基因敲除转化子、互补转化子以及野生型苹果树腐烂病菌菌丝形态和致病力进行分析。与野生型菌株03-8相比,Vmzfp3的敲除转化子的菌丝生长速率明显减慢,致病力也明显下降,其互补转化子使得这些功能缺陷均得以恢复。Vmtme42的敲除转化子无明显区别。由此证明基因Vmzfp3对病菌的菌丝生长,病菌的致病力具有调控作用。
[Abstract]:In this study, the pathogenicity of 150 T-DNA mutants inserted into mutants library of apple tree rot bacteria was screened by inoculating in vitro. After many experiments, three mutants with significantly weakened pathogenicity were obtained, named X4379X 4368C X4468. The results of genome re-sequencing showed that the mutant X4379 had two T-DNA insertion sites, namely zinc finger protein 3 (Vmzfp3) and transmembrane protein 42 (Vmtme 42). The knockout vector and complementary expression vector were constructed based on the whole genome sequence of apple tree rot pathogen. By using PEG method, the knockout transformants and complementary transformants of Vmzfp3 and Vmtme42 were obtained. The mycelium morphology and pathogenicity of Vmzfp3 and Vmtme42 knockout transformants, complementary transformants and wild apple tree rot bacteria were analyzed. Compared with wild-type strain 03-8, the mycelium growth rate and pathogenicity of the knockout transformants of Vmzfp3 were significantly decreased, and their complementary transformants could restore the knockout transformants of .Vmtme42. It was proved that Vmzfp3 could regulate the mycelium growth and pathogenicity of the pathogen.
【作者单位】：河北农业大学植物保护学院;西北农林科技大学植物保护学院/旱区作物逆境生物学国家重点实验室;
【基金】：基金项目:国家苹果现代产业技术体系(CARS-28)
【分类号】：S436.611.11

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