自毒作用是葡萄连作障碍产生的重要原因。为探明自毒物质介导下，葡萄根系分泌物成分的变化机制及其对土壤微生物的作用效应。本项目在前期研究的基础上，采用13CO2脉冲标记技术，将葡萄自毒物质对羟基苯甲酸（浓度覆盖促进—抑制作用）加入到悬空法培养的贝达葡萄幼苗中，检测根缘细胞数量、存活率及黏液层厚度；同时采用LC-MS技术测定根缘细胞黏液层中糖、氨基酸及酚酸含量，从根缘及根尖细胞质膜透性、ROS含量、Ca2+水平以及控制糖、氨基酸及酚酸代谢途径关键酶基因的表达水平上，明确自毒物质对根缘细胞及胞外黏液层成分（根系分泌物的主要来源）的调控机制。同时结合13CO2标记试验，利用稳定同位素核酸探针（RNA-SIP）及Illumina Miseq测序技术，原位鉴定与根缘细胞代谢有关的功能微生物，明确根缘细胞对微生物的调控机制，从根缘细胞和土壤微生物对自毒物质的响应机制角度，揭示自毒物质的作用机理。

Autotoxicity is the important reason of grape replant disease. In order to explore the changing mechanism of root exudate composition and its effect on rhizosiphere soil microorganism under the impact of autotoxicity. Based on previous research, 13CO2 pulse-labeling technique was used in this project and the ρ-hydroxybenzoic acid of grape autotoxin (the concentration from promotion to inhibition) was sprayed to Beta ( V. riparia×V.labrusca) grape seedlings.The number, survival rate and thickness of slime layer were checked, and the sugar, amino acid and phenolic acid contents in slime layer of root border cells were detected by LC-MS technique. In order to expound the regulating mechanism of autotoxin on border cells and extracellular slime layer, which was the main source of root exudate, the cell membrane permeability, ROS content, Ca2+ level and the expression level of key enzyme gene which regulated the metabolic pathways of sugar, amino acid and phenolic acid were measured. In order to clarify the regulating mechanism of border cells on microorganism, the microorganism ralated to border cells was identified by RNA-SIP and Illumina Miseq technique based on 13CO2 pulse-labeling experiment. The aim of our project was to reveal the mechanism of grape autotoxin from the point of border cells and microorganism.