大蒜化感物质消减番茄连作障碍的生理和分子机制

Continuous cropping obstacles are the obvious problems and constraints faced by fruit vegetables production in greenhouse. We have found that the continuous cropping obstacles of tomato can be alleviated by intercropping with garlic, and diallyl disulfide (DADS) identified is the major allelochemical in garlic root exudates and garlic straws, but its physiological and molecular mechanisms are still unclear. In this project we will study the physiological effect of diallyl disulfide on tomato, investigate the allelopathy of diallyl disulfide on tomato root genes expression profiles by RNA-seq, analyse the sensitive metabolic pathways and functions of genes that response to garlic allelochemical and confirm the temporal and spatial expression characteristics and transcription level of major genes by qRT-PCR. We also want to analysze the corelation between the major gene (TAO2) and diallyl disulfide by RNA interference and overexpression technology. The impact of diallyl disulfide on tomato root exudates and especially the autotoxins changes will be studied by GC-MS. Then the growth effect of diallyl disulfide on TAO2 transgenic and non-transgenic tomato plants in the continuous cropping soil will be analysed. These researches will clarify the physiological and molecular mechanism of garlic allelochemical diallyl disulfide in alleviating tomato continuous cropping obstacles, in order to lay the theoretical basis and technological foundation for the development of garlic allelochemicals and the application of physiological and molecular regulation in alleviating fruit vegetable continuous cropping obstacle.?It is of great significance for the safe and sustainable development of fruit vegetables production in greenhouse.

连作障碍是设施果菜生产面临的突出问题和制约因素。我们前期研究发现，套作大蒜可以消减番茄连作障碍，并已从大蒜根系分泌物和秸秆中鉴定出主要化感物质二烯丙基二硫（DADS），但其作用机理尚不清楚。本项目用DADS处理番茄，分析其生理响应；通过RNA-seq测定番茄根系基因表达谱，分析番茄响应大蒜化感物质的敏感代谢途径和主效基因及其功能，通过qRT-PCR分析主要相关基因的时空表达特性；利用RNAi和超表达初步分析主效基因（TAO2）的功能及其与大蒜化感物质响应的关系；通过GC-MS鉴定DADS对番茄根系分泌物成分尤其是自毒物质的影响；比较连作土壤条件下DADS对转TAO2基因和非转基因番茄的生长效应，阐明大蒜化感物质消减番茄连作障碍的生理和分子机制。研究结果将为开发大蒜化感物质及应用生理和分子调控技术消减果菜连作障碍奠定理论依据和技术基础，对设施果菜安全可持生产具有重要意义。

连作障碍已成为我国设施果菜可持续发展的制约因素。课题组前期研究发现，套作大蒜可以消减番茄连作障碍，大蒜根系分泌物的化感作用可能起主要作用。大蒜根系分泌物中主要的化感物质是二烯丙基二硫（DADS），但其作用的生理和分子机制。本项目研究DADS对番茄种子发芽和幼苗生长的生理生化效应、对番茄根系生长和细胞分裂相关基因表达的影响，对番茄敏感代谢途径的调控，筛选与DADS响应有关的主要基因SlDIR1-like，并通过转基因手段分析其作用，分析DADS对连作番茄根系分泌物变化的影响，主要研究结果如下：.1. DADS对番茄的根系生长具有低促高抑的浓度效应现象。.2.较低浓度DADS（0.14和0.21 mM）对番茄幼苗生长有促进作用，增加了植株鲜重、株高、根总长、根体积、根表面积、根系活力和叶绿素含量。DADS可提高番茄籽苗防御酶活性。.3. DADS从3个方面激活番茄的抗性：增加非酶抗氧化物质GSH含量及代谢；诱导抗氧化酶（SOD，POD，CAT，PAL等）活性；激活效应子触发免疫（ETI）抗病途径，诱导系统获得性抗性。.4. DADS通过改变植物激素合成和信号转导途径，影响番茄生长和抗性。.5. DADS能通过调节番茄根系分泌物含量的变化可以缓解番茄自毒作用。.6.SlDIR1-like为DADS主效响应基因之一，可能与系统获得性抗性（SAR）信号形成和胞间传递有关。水杨酸（SA）受体蛋白基因NPR1和SAR的标记基因PR1表达水平上升，说明DADS可能增加了番茄不同组织中SA水平，诱导了SAR反应。构建SlDIR1-like过表达、RNAi和CRISPR敲除载体，进行番茄遗传转化，已获得T1代转基因材料，并继续深入研究该基因的功能。.项目发表署名SCI论文7篇。研究结果为开发大蒜化感物质和生理与分子调控技术消减果菜连作障碍奠定理论依据和技术基础，对设施果菜安全可持续生产具有重要意义。