黄瓜白粉病抗性主效基因的克隆与功能分析

Powdery mildew (PM) is one of the most serious diseases in cucumber production, and breeding of PM resistance cultivars is the most desirable strategy to control the disease. It is very important for resistance breeding to clone the resistant gene and study the molecular mechanism of powdery mildew resistance in cucumber. For the major locus of powdery mildew resistance, with marker-assisted selection, the chromosome segment substitution lines and backcrossing populations would be constructed and used for map-based cloning to isolate this major resistance gene. Then a series of experimental means, such as complementation test of genetic transformation, analysis of tissue-specific gene expression, subcellular localization, transcriptome analysis, yeast two-hybrid system, would be performed to determine the resistance gene and its regulatory network, and to uncover the molecular mechanism of the PM resistance in cucumber. Additionally, for the resistance gene, easy-testing markers would be developed for molecular breeding of PM resistance in cucumber.

白粉病是黄瓜生产中最为严重的病害之一，培育抗白粉病的黄瓜品种是解决其危害的最好方法。白粉病抗性基因的克隆及其分子机制的研究，对黄瓜白粉病抗性育种有重要意义。本项目针对黄瓜白粉病抗性主效基因，通过分子标记辅助选择，构建其染色体片段代换系及其高代回交分离群体，利用图位克隆方法对此抗性主效位点进行克隆。通过遗传转化互补验证、组织表达特异性分析、亚细胞定位、转录组分析、酵母双杂交等实验手段，确定所克隆的抗性基因及其所参与的调控网络，揭示黄瓜白粉病抗性的分子机制。开发抗性基因容易检测的分子标记，用于黄瓜白粉病抗性育种的分子标记辅助选择，加快黄瓜抗病育种的速度。

黄瓜是世界上重要的蔬菜作物之一，白粉病是黄瓜生产中一种普遍的真菌病害，造成了产量与品质的严重损失。本研究构建了黄瓜白粉病抗性自交系S1003的染色体片段代换系及其回交群体，通过图位克隆分离获得黄瓜白粉病抗性主效基因pm5.1，即CsMLO1， 其突变体分为3种类型，分别造成了转录本的缺失、移码突变及提前终止等，使其功能丧失，产生对白粉病的抗性表型，该抗性为隐性。利用S1003及其pm5.1近等基因系NIL(Pm5.1)，探索黄瓜对白粉病的抗性机制。CsMLO1基因受白粉病菌的诱导表达，在接种后12 h表达量最高。qRT-PCR分析显示，近等基因系之间的抗、感差异可能与SA、JA、乙烯途径信号分子无关。转录组分析表明，细胞壁加厚和ABC转运子可能在黄瓜pm5.1所介导的白粉病抗性过程中发挥了很重要的作用。根据CsMLO1基因的插入/缺失差异，成功开发了共显性的InDel标记，可用于白粉病抗性育种的分子标记辅助选择。在pm5.1克隆的基础上，精细定位了另一个抗白粉病基因Pm5.2，有利于揭示黄瓜白粉病抗性机制。