摘要
本论文筛选和鉴定了芽孢杆菌的拮抗菌株，探讨了接种芽孢杆菌提高甘薯对蔓割病菌抗性的光合作用机制和保护酶系统的启动机制，构建了枯草芽孢杆菌绿色荧光蛋白高效表达载体，并从枯草芽孢杆菌168菌株中克隆和表达了抗菌蛋白。结果如下：
(1)以植物根际分离到的41株芽孢杆菌菌株为对象，分别测定了其对植物病原真菌和细菌的拮抗效果，从中筛选出11株对植物病原菌抑制率较高的菌株，即AB、V3、SAI、3-1、N8-b、25-1、25-5、25-6、25-9、25-15、25-17。并分别研究了对甘薯蔓割病菌(Fusarium．oxysporumf．sp．batatas)、小白菜黑斑病菌(Alternaria brassicae)、香蕉炭疽病菌(Colletotrichum musae)、立枯丝核菌(Rhizoctonia soIani)、辣椒疫霉病菌(Phvtophthora capsici)的抑菌作用，其中着重研究了菌株25-5、25-6、25-9、25-15、25-17对甘薯蔓割病菌、小白菜黑斑病菌，菌株25-1、25-17对香蕉炭疽病菌的作用机理。结果表明：平板对峙培养出现明显的抑菌环；以凹玻片法，用菌株培养滤液处理病原菌分生孢子，其孢子萌发率明显降低；将细菌培养滤液和孢子悬浮液涂布于PDA平板上，培养滤液均能抑制病菌分生孢子的形成。普通光学显微镜和油镜观察显示，菌株均可抑制病菌菌丝生长，造成菌丝分支增多，顶端和中央膨大，随着时间的延长畸变结构增加，菌丝呈念珠状或菌丝分支显著增多，且顶端细胞变形膨大聚集成簇，菌丝壁穿洞，内含物质泄漏，终使细胞崩解和消融，变成球状的空泡。
(2)以芽孢杆菌菌株25-5、25-6、25-9、25-15、25-17为材料，探讨了接种芽孢杆菌提高甘薯对蔓割病菌抗性的光合作用机制和保护酶系统启动机制，结果表明：接种芽孢杆菌后的甘薯叶片光合速率、气孔导度、蒸腾速率均明显高于对照，这些作用随蔓割病病原菌的浓度升高而下降，处理间表现一致；同时降低了MDA含量，提高植株的活性氧保护酶系统代谢水平。但是经培养基处理的甘薯叶片光合速率、活性氧保护酶系统代谢水平却反而下降，说明接种芽孢杆菌可以有效提高甘薯的光合作用，增强抵御蔓割病菌的能力。
(3)构建了枯草芽孢杆菌绿色荧光蛋白高效表达载体，将枯草芽孢杆菌组成型启动子P43克隆到枯草芽孢杆菌—大肠杆菌穿梭载体pGFP4412中，构建了携带绿色荧光蛋白的重组质粒pP43GFP，结果发现枯草芽孢杆菌组成型启动子P43启动绿色荧光蛋白基因在枯草芽孢杆菌和大肠杆菌表达，成功地对枯草芽孢杆菌模式菌株BS168进行了标记，为进一步用gfp基因标记植病生防枯草芽孢杆菌和外源蛋白表达打下技术基础。
(4)利用大肠杆菌——枯草芽孢杆菌穿梭表达载体P43GFP将TasA基因与枯草芽
This dissertation studied the screening and identification of antagonistic strains of Bacillus spp., and discussed the mechnisms of Bacillus spp. to improve sweet patato' s resistance to Fusarium. oxysporumf. sp. batatas., which included increasing photosynthesis and promoting protective enzyme system. The main results were as follows:
(1) Forty-one strains of Bacillus spp. were obtained from plant rhizosphere and their antagonistic effect on plant fungal and bacterial pathogens were tested. Among them, eleven strains with high inhibitory rate on plant pathogen were screened, which were AB, V3, SAI, 3-1, N8-b, 25-1, 25-5, 25-6, 25-9, 25-15 and 25-17. This research also explored the inhibition mechanism of Bacillus spp. on several pathogens, which were F. oxysporumf. sp. batatas, Alternaria brassicae, Colletotrichum musae, Rhizoctonia solani, Phytophthora capsici. The research of mechanism focused on inhibitory effects of strains 25-5, 25-6, 25-9, 25-15, 25-17 on F. oxysporum f. sp. batatas, Alternaria brassicae and those of strains 25-1, 25-17 on Colletotrichum musae. The results indicated as follows: with dual culture on plate method, significant anti-bacterial circle was observed; with concave slide method, in which the pathogen conidia were treated with bacterial culture filtrate, the germination rate of conidia decreased significantly. Coating bacterial culture filtrate and conidia suspension on the PDA plate, culture filtrate inhibited pathogen conidia' s formation. Observing strains with optical microscope and optical oil immersion lens showed that all of the strains could inhibit pathogen growth. The inhibition caused the increase of pathogen hyphae branches and the expansion of the top and central part. With continued inhibition, abnormal structures increased, the hypha became bead-like or with more branches, and the cells on the end of the hypha expensed to clusters, the hypha walls pierced so the contained substance leeked out, the cells collapsed and turned into hollow sphere bubble.
(2)With Bacillus spp. strains 25-5, 25-6, 25-9, 25-15, 25-17 as materials, the photosynthetic mechanism and starting mechanism of protective enzyme system in the process of inoculating Baccillus spp. to increase sweet potato' s resistance to F. oxysporumf. sp. batatas, were investigated. The results indicated that for the sweet potato leaves treated with Baccillus spp.
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