MOLECULAR ANALYSIS OF MYCOBACTERIAL GENE EXPRESSION

分枝杆菌基因表达的分子分析

• 批准号: 2671372
• 负责人: Gerard J Nau
• 金额: $ 8.88万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-01 至 2000-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2671372
• 关键词: Mycobacterium; Mycobacterium tuberculosis; affinity chromatography; bacterial genetics; cellular immunity; gene expression; genetic library; genetic promoter element; genetic transcription; operon; phagocytosis; reporter genes; site directed mutagenesis; tissue /cell culture; transfection /expression vector; vector vaccine; virulence

The proposed project will examine the adaptive responses m. tuberculosis undergoes after phagocytosis by a macrophage. Genes associated with infection are likely to be regulated and are likely to be under the control of specific promoters responsive to environmental cues. The main objective of the project will be to assess activation and inactivation of gene transcription by analysis of promoter activity; a reporter vector system will be developed for this purpose. Genomic M. tuberculosis DNA will be fragmented and ligated into the reporter vector, which will then be used to transform M. tuberculosis. Transformants will be subjected to phagocytosis by a macrophage cell line; promoters with altered activity will be identified by differential expression of the reporter genes. Sequences that flank these promoters will be assessed for protein coding sequences and regulatory regions. Genes identified from these sequences will be evaluated for their relevance to virulence, which will be tested ultimately by site-specific mutagenesis of wild-type M. tuberculosis. The proposed work will provide fundamental information regarding the structure and regulation of mycobacterial promoters, genes, and operans. In addition, understanding M. tuberculosis virulence factors could identify new targets for vaccine or drug development.

拟议的项目将检查自适应响应m。结核 巨噬细胞在吞噬吞噬作用后经历。 与之相关的基因 感染可能受到调节，并可能受到控制 对环境提示有效的特定启动子。 主要目标 该项目将是评估基因的激活和失活 通过分析启动子活性转录；记者向量系统 将为这个目的开发。 基因组分枝杆菌DNA将是 分散并连接到报告基因向量，然后将其用于 转化结核分枝杆菌。 转化子将受到吞噬作用 通过巨噬细胞系；活动变化的发起人将是 通过报告基因的差异表达鉴定。 序列 这些启动子将用于蛋白质编码序列的侧面 和监管区域。 从这些序列中确定的基因将是 评估它们与毒力的相关性，最终将进行测试 通过野生型结核病的位点特异性诱变。 提议 工作将提供有关结构和 分枝杆菌启动子，基因和操作剂的调节。 此外， 了解结核病毒力因素可以识别新目标 用于疫苗或药物开发。