EXITING STATIONARY PHASE IN E COLI

退出大肠杆菌中的固定相

• 批准号: 6417963
• 负责人: DEBORAH A. SIEGELE
• 金额: $ 6.09万
• 依托单位: TEXAS A&M UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-05-01 至 2003-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6417963
• 关键词: Escherichia coli; bacterial genetics; bacterial proteins; gene expression; gene mutation; mass spectrometry; microorganism growth; polymerase chain reaction; protein sequence

DESCRIPTION: Bacterial infections remain a leading cause of illness and death. To control bacterial pathogens it is important to understand not only host-pathogen interactions, but also how the pathogen's life cycle is affected by the environment. Bacteria can survive a variety of environmental stresses and still rapidly resume growth when conditions become favorable. This ability is an important factor in disease transmission and can also affect the initial stages of infection. The long-term goal of this project is to understand the mechanisms that regulate gene expression as the enteric bacterium E coli recovers from starvation stress. Mechanisms we identify are likely to be important in the infection cycle of pathogenic E. coli and other pathogenic bacteria. A group of nine E. coli proteins are transiently expressed during the initial period of outgrowth from stationary phase. The limited time when these proteins are made suggests that they are controlled by factors involved in regulating important aspects of the outgrowth response. The immediate goal of this proposal is to identify the genes encoding outgrowth-specific proteins. Once identified, these genes will be used as reporters to study the regulation of the outgrowth response. The first specific aim is to identify genes encoding outgrowth-specific proteins by obtaining amino acid sequence information from the outgrowth-specific proteins already described. State-of-the-art mass spectrometry methods will be used to analyze gel-purified proteins. The sequence information obtained will be used to identify the genes in the completed E. coli genome sequence. The second aim is to identify transcripts made specifically during outgrowth using PCR based differential display. The sequence of the differentially expressed cDNAs will be compared to the E. coli genome sequence to identify the genes. Once outgrowth-specific genes are identified, their function. and the mechanisms regulating their expression will be studied. Mutations will be constructed and the phenotypes of mutants characterized. Standard molecular methods, such as Northern blots and fusions to reporter genes, will be used to characterize how expression is the outgrowth-specific genes is controlled.

描述：细菌感染仍然是疾病的主要原因， 死亡 为了控制细菌病原体，重要的是要了解， 不仅包括宿主与病原体的相互作用，还包括病原体的生命周期 受到环境的影响。 细菌可以在多种环境中存活 环境压力，当条件允许时， 变得有利。 这种能力是疾病的重要因素 传播，也可以影响感染的初始阶段。 的 这个项目的长期目标是了解调节的机制， 大肠杆菌从饥饿中恢复时的基因表达 应力 我们确定的机制可能在感染中很重要 致病性E.大肠杆菌等致病菌。 一行九 E.大肠杆菌蛋白质是瞬时表达的初始阶段， 稳定期的产物。 当这些蛋白质被 提出，它们是由参与调节的因素控制的， 生长反应的重要方面 这件事的直接目标是 这项研究的目的是鉴定编码生长特异性蛋白质的基因。 一旦确定，这些基因将被用作报告基因， 调节生长反应。 第一个具体的目标是确定基因编码的生长特异性 蛋白质的氨基酸序列信息， 已经描述的生长特异性蛋白质。 最先进的质量 光谱法将用于分析凝胶纯化的蛋白质。 的 所获得的序列信息将用于鉴定 完成E。coli基因组序列。 第二个目标是确定 使用基于PCR的差异表达技术， 显示. 差异表达的cDNA的序列将被 与E.大肠杆菌基因组序列，以确定基因。 一旦确定了生长特异性基因，它们的功能。 和 将研究调节其表达的机制。 突变将被 构建并表征突变体的表型。 标准分子 将使用诸如北方印迹和与报告基因融合的方法 来描述特定基因的表达是如何产生的， 控制。