GFP标记内生枯草芽孢杆菌Y10及其在白菜体内的定殖  

Colonization of GFP-tagged endophytic Bacillus subtilis Y10 in Chinese cabbage

作　　者：杜芳[1] 何鹏飞[1] 吴毅歆[2,3] 陈卓君[1] 杨静[1] 何月秋[2,3] 

机构地区：[1]云南农业大学植物保护学院,昆明650201 [2]微生物菌种筛选与应用国家地方联合工程研究中心,昆明650207 [3]云南农业大学农学与生物技术学院,昆明650201

出　　处：《生态学杂志》2015年第7期2064-2070,共7页Chinese Journal of Ecology

基　　金：农业部公益性行业(农业)专项(201003029)资助

摘　　要：枯草芽孢杆菌Y10是具有防治根肿病的白菜内生菌株。为研究其在白菜体内的定殖规律,将不同来源的含绿色荧光蛋白（GFP）质粒导入Y10进行荧光标记,测定了荧光标记菌的质粒稳定性、生长曲线及室内平板拮抗活性。结果表明,仅有p HT01-P43GFPmut3a能在Y10中稳定地遗传,且对Y10的生长和病原真菌拮抗活性无不利影响。盆栽防效试验结果显示,在不同的栽培基质中,Y10的标记菌株与野生型对白菜根肿病防效无显著差异。稀释涂板回收及共聚焦激光扫描显微观察都证实,接种9 d后在白菜的根、茎及叶部组织均有标记菌株的定殖,且在44 d内仍保持103~104cfu·g-1组织。这些定殖结果表明,Y10在白菜体内具有良好的定殖能力,且这种特性可能与其防治根肿病有关。Endophytic Bacillus subtilis YI（）isolated from Chinese cabbage （Brassica rapa） can control effectively cruciferous vegetable clubroot disease. To understand its clubroot-control mech- anisms, GFP plasmids were transformed into Y10 cells for fluorescence-tagging by natural trans- formation. The plasmid stability, growth curve and plate antagonism bio-assay of GFP-tagged strains were determined, and the results showed only plasmid pHT01-P43GFPmut3a could be maintained stably and its introduction had no significant negative influence on ~10 growth and antagonistic performance. Pot bio-assay experiment with different cultivation matrixes indicated that there was no significant difference in clubroot-control efficiency between Y10-P43GFPmut3a and its wi｜d type. Recovery and observation by confocal laser scanning microscope confirmed that, 9 days after inoculation, tagged strain could colonize the root, stem and leaf of Chinese cabbage, and the colonization density still kept at 103-104 cfu · g-1 tissues in 44 days after inoc- ulation. All the results suggest that the clubroot-control of Y10 be intimately associated with effi- cient colonization in Chinese cabbage host.

关 键 词：内生枯草芽孢杆菌 绿色荧光蛋白 防治效果 质粒 遗传转化 

分 类 号：S476.1[农业科学—农业昆虫与害虫防治]