菵草（Beckmannia syzigachne）对精噁唑禾草灵和甲基二磺隆多抗性的分子机制

Along with the application of weed-killer, some weeds have evolved resistance to herbicide. Applying herbicides scientifically on the base of resistant mechanisms is the only way to control resistant weeds effectively. Evolved target-site resistance (TSR) and non-target-site resistance (NSTR) are both found in the herbicide-resistant weeds. The TSR mechanism consists of an amino acid change in a target enzyme conferred by gene mutation and an overexpression of a target enzyme gene. NSTR mechanism includes an increased herbicide metabolism, an increased rate of herbicide translocation, and decreased rates of herbicide translocation. Different mechanisms endow weeds different resistant index and different cross resistance. American sloughgrass (Bechmania syzigachne) is one of the main weeds in wheat fields rotated with rice. Either fenoxaprop-p-ethyl or mesosulfuron is important herbicide applied in wheat fields to control American sloughgrass. But American sloughgrass have evolved multiple-resistance to fenoxaprop-p-ethyl and mesosulfuron in fields. These multiple-resistant populations are great threat to wheat production. These multiple-resistant American sloughgrass populations can't be controlled effectively because of the inadequate information about resistant mechanisms. We will take the multiple-resistant American sloughgrass populations as target in this project. Biochemistry methods and molecular biology methods will be employed to research the molecular resistance mechanism. With completion of this project, the molecular mechanism of multiple-resistance to fenoxaprop-p-ethyl and mesosulfuron in American sloughgrass will be determined a rapid detection method will be built based on the molecular mechanism.

随着除草剂使用历史的增长，部分杂草对除草剂产生了抗药性，针对其抗药性机制合理选用除草剂是防除抗药性杂草的有效途径。杂草对除草剂产生抗药性的机制主要包括靶标酶基因突变导致的氨基酸取代、靶标酶基因表达量增加、对除草剂代谢能力的增强以及对除草剂吸收转运的减少，不同的机制会赋予杂草不同的抗药性水平和交互抗性。 菵草是稻茬麦田的主要杂草，危害严重，主要依靠精噁唑禾草灵和甲基二磺隆防治。对精噁唑禾草灵和甲基二磺隆同时产生抗性的菵草种群已在田间发生，构成对农业生产的巨大威胁。由于目前对其抗药性机制所知甚少，难以进行有效的防治。 本项目拟以对精噁唑禾草灵和甲基二磺隆产生多抗性的菵草为研究对象，从分子水平研究其抗药性的分子机制，以期明确其对精噁唑禾草灵和甲基二磺隆产生多抗性的分子机制，并利用CAPs（dCAPs）技术建立一种快速检测方法。

对精噁唑禾草灵和甲基二磺隆产生多抗性的菵草是稻麦轮作区小麦生产的严重威胁，为了明确多抗性菵草对精噁唑禾草灵和甲基二磺隆的分子抗性机理，本项目以对精噁唑禾草灵和甲基二磺隆产生多抗性的鱼台种群和丹阳种群为研究对象，测定了其抗性水平、对其他药剂的交互抗性，研究了菵草对精噁唑禾草灵和甲基二磺隆多抗性的分子机理，测定抗性突变位点基因的纯合性，研究结果表明：.1. 丹阳种群和鱼台种群高抗精噁唑禾草灵，抗性指数分别为565.0和236.8；对甲基二磺隆有中等水平抗性，抗性指数分别为6.7和6.4。丹阳种群对炔草酯、唑啉草酯、烯草酮存在交互抗性，烯禾啶可作为替代药剂；鱼台种群对炔草酯、烯禾啶、烯草酮有交互抗性，唑啉草酯防效良好。丹阳种群和鱼台种群对啶磺草胺、氟唑磺隆都有交互抗性。抗性种群对异丙隆均无抗性。.2.抗性分子机理研究表明丹阳种群ACCase的Trp1999Leu取代和鱼台种群ACCase的Gly2096Ala取代是其对精噁唑禾草灵产生抗性的分子机理；丹阳种群ALS的Pro197Ser取代是其对甲基二磺隆产生抗性的分子机理，鱼台种群ALS基因序列中不存在已知的抗性突变位点。.3. (d)CAPS分析表明鱼台种群ACCase 2096位点、丹阳种群ACCase 1999位点均为杂合基因，其中丹阳种群纯合子比例更高。PCR产物测序表明丹阳种群ALS 197位点为杂合基因，但抗性植株均为突变纯合子。.4. RT-q-PCR分析发现抗性种群ACCase基因表达量与敏感种群无显著差异，转录组测序数据表明鱼台种群ALS基因表达水平与敏感种群无显著差异。.5. HPLC分析发现鱼台种群野生型代谢精噁唑禾草酸的速度显著快于敏感生物型。对鱼台种群和泰安种群进行转录组测序，获得了115112条Unigene，并进行了注释。对差异表达基因进行分析发现，敏感种群与抗性种群存在诸多差异表达基因，其中包含P450、GST等代谢相关基因。.本项目的研究结果明确了菵草对精噁唑禾草灵和甲基二磺隆多抗性的分子机理，丰富了对杂草抗性的认识，筛选了防治药剂，对多抗性菵草的防治具有很好的指导意义。