【作者】 杜红方；

【导师】 张元亮；

【作者基本信息】 河北大学 ， 植物学， 2003， 硕士

【摘要】 本文在对棉花生态系统中的细菌进行了全面的分离的基础上，通过体外拮抗试验筛选出一株对棉花黄萎病具有拮抗作用的芽孢杆菌LC105菌株，对其进行了菌种的初步鉴定和产抗菌蛋白发酵条件研究，并从其发酵液中分离纯化出单一组分的抗棉花黄萎病菌的蛋白质。 从棉花各部位共分离出芽孢菌148株，其中有38株表现出对棉花黄萎病大丽轮枝菌的拮抗活性，其中有一株编号为LC105从棉花叶部分离出的芽孢杆菌表现出很强的拮抗活性。 对编号为LC105的菌株进行了形态观察和生理生化的实验研究初步认定该芽孢菌属类芽孢杆菌属。对其发酵液进行硫酸铵盐析，发现沉淀物的溶液对棉花黄萎病具有拮抗作用；对其进行加热处理和有机溶剂(氯仿)处理，发现其抗菌活性消失，初步认定其拮抗物质为蛋白质。 采用不同的培养温度、不同的培养时间、不同的摇床转速、不同的培养基和不同的pH值对拮抗菌LC105进行培养，对其发酵液的抗菌活性进行检测，得到了抗菌蛋白产生的最佳条件为：在pH值为7的LBG培养基中30℃、转速180r／min摇床振荡培养48小时，以50～70％饱和度的硫酸铵对其进行分级盐析，可得最大量的抗菌蛋白。 通过硫酸铵的分级沉淀、DEAE-Sephadex A-50离子交换层析和Sepharyl S-100分子筛凝胶层析的方法，从LC105菌株的发酵液中得到了一种对棉花黄萎病菌具有拮抗作用的抗菌蛋白。更多还原

【Abstract】 This paper presents a study on the biological control of cotton disease Verticillium wilt by antagonistic bacteria including the isolation, screening of antagonistic bacteria and purification for an antifungal protein.148 strains bacteria were isolated from cotton plant microecosystem. The antagonistic assay in plates showed that among them there were 38 strains which had antagonistic activity to Verticillium dahliae. LC105 had the highest antagonistic activity.Proteins were precipitated from ferment solid production of LC105 and they had the antagonistic activity to Verticillium dahliae. The antagonistic activity of the proteins were lost after were treated by heat and CHCl3. This showed that the antifungal material must be protein.We cultured LC105 strain under different conditions and studied the antagonistic activities of the solid ferment production. The assay showed that the strain could serecte lots of antifungal protein under 30℃ degree, in LBG medium of pH 7 and cultured 48 hours.With (NH4)2S04 fractional precipitation, DEAE-Sephadex A-50 ion exchange chromatography and Sephacryl S-300 molecular filtration chromatography followed by tracks of fraction antagonistic assay and PAGE, an antifungal protein with in vitro antagonistic activity to Verticillium dahliae was isolated and purified.更多还原