METHOD FOR ISOLATING AND STUDYING MARINE MICROORGANISMS

分离和研究海洋微生物的方法

• 批准号: 2866839
• 负责人: YEVGENYA AKSELBAND
• 金额: $ 11.46万
• 依托单位: ONE CELL SYSTEMS, INC
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-30 至 2000-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2866839
• 关键词: aquatic organism; bacteria characteristic; flow cytometry; fluid; fluorescence; gel; growth media; marine biology; microorganism culture; microorganism metabolism; water environment

Marine microorganisms are a rich potential source of novel metabolites with unique biotherapeutic applications. Conventional isolation methods include plating on liquid selective enrichment media or specially formulated solid media. However, only about 5% of these microorganisms can be isolated because many fail to form colonies on solid media. Viable slow growing or non-growing organisms, which comprise up to 80% of some collections, are also quickly overgrown by fast growers using liquid culture media. To date, isolation based on metabolic activity, especially in the case of bacteria smaller than 5 mu m, has not been possible. This Phase I proposal aims to develop a rapid, high throughput, automated assay to analyze and isolate bacteria from environmental samples based on their biochemical properties, and metabolic and proliferative activities. Using phytoplankton as a model system, a gel microdrop (GMD) assay which permits flow cytometric sorting of different bacteria based on autofluorescence will be developed. To isolate bacteria based on proliferative activity, GMD encapsulation and fluorescence activated cell sorting (FACS) will be combined. To isolate bacteria based on metabolic activity, GMD encapsulation, 5-cyano-2,3-ditolyl tetrazolium chloride(CTC) which detects cellular respiratory activity, and FACS will be combined. PROPOSED COMMERCIAL APPLICATIONS: The proposed assays will facilitate drug discovery by increasing the number of isolated strains of marine microorganisms available for drug screening.

海洋微生物是具有独特的生物治疗应用的新型代谢产物的丰富潜在来源。常规的隔离方法包括在液体选择性富集培养基或专门配制的固体培养基上铺板。但是，这些微生物中只有大约5％可以隔离，因为许多微生物未能在固体培养基上形成菌落。使用液体培养基的快速种植者迅速生长了可行的缓慢生长或非生长生物，最多占某些收藏的生长生物。 迄今为止，基于代谢活性的分离，尤其是在细菌小于5 mu m的情况下，是不可能的。该阶段I建议旨在开发一种快速，高通量，自动化的测定法，以根据其生化特性以及代谢和增殖活性来分析和分离环境样品。将使用浮游植物作为模型系统，将开发基于自动荧光的不同细菌的流式细胞术分类的凝胶微旋转（GMD）测定。为了基于增殖活性分离细菌，将组合GMD封装和荧光激活的细胞分选（FACS）。为了基于代谢活性，GMD封装，5-甲状-2,3-二丙基四唑烷氯化物（CTC）分离细菌，该细菌将检测到细胞呼吸活性，FACS将合并。拟议的商业应用：拟议的测定法将通过增加可用于药物筛查的海洋微生物菌株的数量来促进药物发现。