应用启动子诱捕策略筛选并初步鉴定肺炎链球菌宿主体内诱导基因

发布时间：2018-04-27 07:42

本文选题：肺炎链球菌 + 体内诱导基因　；参考：《重庆医科大学》2005年博士论文

【摘要】：肺炎链球菌(Streptococcus pneumoniae,S.pn)是一种常见的革兰阳性条件致病菌,可引起肺炎、中耳炎、菌血症、脑膜炎等严重疾病,在全球有很高的发病率和死亡率。老人、儿童以及艾滋病和先天免疫缺陷的患者是严重 S.pn 感染的高危人群。由于 S.pn 抗生素耐药率的增加和目前疫苗的缺陷性,使得其感染的后果广泛、严重。因此要想从根本上解决 S.pn 的防治问题,就要深入了解其致病的分子机制,为研发新的药物和疫苗提供理论和实验依据。病原菌侵入宿主体内后,就进入了一个生存竞争的“战场”,为了自身的生存及应对宿主免疫系统的攻击它将调整其基因表达的模式,下调一些非必需基因的表达,上调另一些基因的表达,这些上调的基因是病原菌在宿主体内生存所必需的,它们很可能是致病过程中的关键基因,称之为体内诱导基因(in vivo-induced gene, ivi gene),是潜在的抗生素作用靶点和疫苗候选者。本研究应用基于启动子诱捕策略的体内表达技术(in vivo expression technology,IVET),以肺炎链球菌荚膜合成的关键基因 galU 作为体内报告基因,利用其缺陷体不能合成荚膜多糖,从而不能在宿主体内存活的特点,筛选鉴定肺炎链球菌 ivi 基因。研究内容包括以下三个部分: 1、构建肺炎链球菌荚膜合成的关键基因 galU 缺失的突变体,获得肺炎链球菌荚膜缺陷菌株,并研究其生物学功能,为后续肺炎链球菌体内诱导基因的鉴定提供实验菌株和技术平台。采用长臂同源多聚酶链式反应(Long flanking homology polymerase chain reaction,LFH-PCR)方法,制备中间为红霉素耐药基因(erm),
[Abstract]:Streptococcus pneumoniae S.pn) is a common gram-positive opportunistic pathogen, which can cause severe diseases such as pneumonia, otitis media, bacteremia, meningitis and so on. The elderly, children and people with AIDS and congenital immunodeficiency are at high risk of severe S.pn infection. Because of the increase of antibiotic resistance rate of S.pn and the defect of current vaccine, the result of infection is widespread and serious. Therefore, in order to solve the problem of prevention and cure of S.pn, it is necessary to deeply understand the molecular mechanism of its pathogenesis and provide theoretical and experimental basis for the development of new drugs and vaccines. After invading the host body, the pathogen enters a "battlefield" of survival competition. In order to survive and respond to the attack of the host's immune system, it will adjust its gene expression pattern and down-regulate the expression of some non-essential genes. Upregulate the expression of other genes that are necessary for the pathogen to survive in the host, and that are likely to be key genes in the pathogenesis. Called in vivo-induced gene, ivi gene is a potential antibiotic target and vaccine candidate. In this study, the in vivo expression technique based on promoter trapping strategy was used to study the expression of galU, the key gene of capsule synthesis of Streptococcus pneumoniae, which could not synthesize capsular polysaccharides by using its defective body. The ivi gene of Streptococcus pneumoniae was screened and identified. The research includes the following three parts: 1. To construct the mutant of galU deletion, which is the key gene in the capsule synthesis of Streptococcus pneumoniae, to obtain the strain with deficient capsule of Streptococcus pneumoniae, and to study its biological function. To provide experimental strains and technical platform for identification of induced genes of Streptococcus pneumoniae in vivo. The long arm homologous polymerase chain reaction (long flanking homology polymerase chain reactionn LFH-PCR) was used to prepare erythromycin resistance gene (Erythromycin).
【学位授予单位】：重庆医科大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R378

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