Stabilization of Recombinant Plasmids in Bacillus Subtilis

枯草芽孢杆菌中重组质粒的稳定性

• 批准号: 8711725
• 负责人: Arnold Demain
• 金额: $ 50.72万
• 依托单位: Massachusetts Institute of Technology
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-08-01 至 1991-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=8711725&HistoricalAwards=false
• 关键词: Stabilization; Recombinant; Plasmids; Bacillus; Subtilis

The stability of recombinant DNA (plasmid) in bacteria used for commercial scale fermentations is of great concern. Bacillus subtilis is a potentially useful microorganism for the production of recombinant DNA products. However, instability of plasmids in B. subtilis has limited its usefulness as an industrial cloning host. The overall objective of this study is to analyze genetic, nutritional and environmental factors that affect the stability of recombinant DNA in B. subtilis. The PIs propose to quantitate the parameters that govern the kinetics of plasmid loss, i.e., to measure - The frequency with which segregational events or structural changes in the plasmid occur. - The growth advantage the modified cells have over parental cells. Chemostat studies will be employed as a unique tool for delineating the relative effects of segregational and structural plasmid instabilities. Using state-of-the-art fermentor monitoring and estimation methodologies, the on-line detection of revertants in the recombinant culture will be attempted at an early stage to allow for the application of stabilization strategies in fed-batch and continuous culture.

重组DNA（质粒）在细菌中的稳定性 商业规模的发酵受到极大关注。 芽孢杆菌 枯草杆菌是一种潜在的有用的微生物，用于生产 重组DNA产物。 而质粒在B. 枯草芽孢杆菌作为工业克隆的用途有限 主持人 这项研究的总体目标是分析遗传， 营养和环境因素影响的稳定性 重组DNA在B中。枯草杆菌。 PI建议量化控制 质粒丢失的动力学，即，测量 - 分离事件或 质粒发生结构变化。 - 修饰细胞的生长优势超过 亲代细胞。 恒化器研究将作为一种独特的工具， 描述分离和结构的相对影响 质粒不稳定性。 使用最先进的发酵罐 监测和估计方法，在线检测 将尝试重组培养物中的回复突变体， 早期阶段，以便在 补料分批和连续培养。