3种药用植物内生真菌的多样性分析及产生物碱菌株的筛选  

Biodiversity of endophytic fungi and screening of alkaloid-producing fungi from three kinds of medicinal plants

作　　者：杨迎迎[1,2] 熊智强[2] 王勇[2] 

机构地区：[1]华东理工大学生物工程学院生物反应器国家重点实验室,上海200237 [2]中国科学院上海生命科学研究院植物生理生态研究所合成生物学重点实验室,上海200032

出　　处：《中国抗生素杂志》2013年第12期901-908,共8页Chinese Journal of Antibiotics

基　　金：国家重点基础研究发展计划(973)项目(2012AA02A701);国家自然科学基金项目(31170101&31100073);中科院知识创新项目(KSCX2-EW-J-12)

摘　　要：目的探究药用植物蛇足石杉、曼地亚红豆杉和南方红豆杉内生真菌的多样性,以及从中筛选产生物碱的内生真菌。方法用传统的表面灭菌和组织剪切法处理植物样品,通过加有抗细菌抗生素的真菌培养基分离和富集植物内生真菌。根据内生真菌的表型特征和ITS序列,对其进行分类和多样性分析,并用Neighbour-Joining的方法构建系统进化树。利用Dragendorff试验检测各内生真菌的发酵液,筛选具有产生物碱潜力的菌株。结果从3种植物中共分离到764株内生真菌,挑选出具有代表性表型特征的菌株83株,对ITS序列进行比对和多样性分析表明,这些菌株分布于子囊菌门(Ascomycota)的3个纲、18个属。最后,采用Dragendorff试验从这83株真菌中筛选出青霉菌属(Penicillium)B14菌株具有产生物碱的潜力。讨论三种药用植物含有丰富多样的内生真菌分布。采用Dragendorff试验检测发酵液中的生物碱,该方法快速简便,能有效提高筛选速率,可应用于产生物碱微生物的高通量筛选。Objective Biodiversity of endophytic fungi and screening of alkaloid-producing fungi were investigated in three medicinal plants, Huperzia serrata, Taxus x madia and Taxus chinensis var. mairei. Methods The plant samples were treated by traditional surface sterilization method and imprinted onto the selective medium for isolation of endophtic fungi. The endophytic fungi were classified based on their phenotypic characteristics and ITS sequences. Phylogenic tree was constructed using the Neighbour-Joining method. Dragendorff assay was employed to screen alkaloid-producing fungi. Results Totally, 763 endophytic fungi were isolated, among which 83 isolates with typical phenotype were chosen to analyze the biodiversity based on their ITS sequences. These isolates were grouped into 18 genera from 3 classes of Ascomycota. Among them, strain B14, belonging to the genus Penicillium, was proved to have potential ability of producing alkaloid by dragendorff assay. Conclusion There are rich and diverse endophytic fungi in these three medicinal plants. Dragendorff assay was a simple and quick method to detect alkaloid from fermentation broth, it thus could be used for high throughput screening of alkaloid-producing microbes.

关 键 词：药用植物 内生真菌 多样性 生物碱 

分 类 号：R978[医药卫生—药品]