枯草芽孢杆菌多胺合成途径产生的促生物质鉴定及其促生机理研究

芽孢杆菌促进植物生长和防治植物病害机制的研究是其高效利用的基础。本课题组前期研究发现，枯草芽孢杆菌OKB105菌株具有很好的促生活性，而突变其多胺合成相关基因yecA和speB后，该菌株促生活性丧失。因此，我们推测这种促生物质来自多胺合成途径。迄今，尚未见类似的报道。本项目将突变多胺合成途径中的8个基因和6个调控基因，再进行功能恢复，对这些菌株代谢产物进行组份分析与分离纯化，并利用核磁共振等技术检测该物质的分子量与结构，从而鉴定该物质及其合成机理。同时，本项目将采用基因标记和凝胶阻滞电泳（EMSA）等方法和技术，解析芽孢杆菌调控该促生物质合成的机理。本项目还将采用Real time PCR和Elisa等技术，检测植物体生长相关基因的表达以及激素水平变化等，研究该物质的促生机制。本项目能揭示芽孢杆菌与植物互作的新机制，并为采用基因工程手段提高芽孢杆菌的生防效果提供可利用的基因。

The researches on the mechanisms of the growth promotion and biocontrol disease are the basis for efficient use of Bacillus spp. In our previous study, we demonstrated that Bacillus subtilis OKB105 could significantly promote plant root growth. But the mutation of yecA and speB, polyamine synthesis-related genes, can disrupt the growth-promoting activity of OKB105. Base on this result, we speculate that substance related to growth-promoting activity is derived from polyamines biosynthetic pathways. Till now, there is no publication related to this. This project will focus on the identification of promoting substances by site-directed mutagenesis, functional recovery, analysis and purification of the metabolite composition, nuclear magnetic resonance and other techniques and methods. At the same time, gene targeting and EMSA would be used to study the regulatory mechanism of this substance. To study the mechanism of action of the substance, Realtime PCR and Elisa technology will be used to detect the expression level of growth-related genes of plant and the level of plant hormones. This project would find out the new mechanism of interaction between plant and B. subtilis, and provide available genes for improving its biocontrol effect by gene engineering.