摘要：阿尔兹海默症是最普遍的一种痴呆症，随着社会老龄化现象的加重，该病的发病率 也随之逐渐提升。许多原因会导致该病的发生，其中大脑中淀粉样多肽 Aβ水平上升及 Aβ错误折叠被普遍认为与阿尔兹海默症的发生直接相关。研究表明，Aβ可以错误折叠为 富含β折叠结构的纤维状聚集体，这些聚集体具有神经细胞毒性，对人体造成伤害。本研 究目的是采用基因重组技术，通过大肠杆菌原核表达出β-淀粉样蛋白。实验方法为根据已 有的 Aβ40 和 Aβ42 基因模板和引物，通过 PCR 反应扩增出 Aβ42 基因片段，定向克隆到载 体质粒 pET-28a 中，将成功构建的重组体 pET-28a-Aβ42 转化到大肠杆菌 DH5α，经过限制 酶双酶切鉴定和 DNA 测序之后，确定基因重组表达载体的成功构建。将筛选的阳性克隆 体通过 IPTG 诱导表达，经 SDS-PAGE 电泳显示表达了β-淀粉样蛋白。本课题成功克隆了 目的基因 Aβ40 和 Aβ42，而且在大肠杆菌中获得了稳定表达。为生产重组β-淀粉样蛋白以 及对阿尔茨海默病的发病机制和临床研究提供了物质和技术保障，为 Aβ的结构功能及动 态过程的研究提供支持。
关键词 阿尔兹海默症 β-淀粉样蛋白 基因表达 克隆
毕 业 设 计 说 明 书 外 文 摘 要
Title Chemical synthesis and biological expression of peptides from Alzheimer's disease model
Abstract:Alzheimer's disease is the most common type of dementia, with the phenomenon of social aging is deepening, the incidence rate is gradually increasing. Many aspects will lead to the occurrence of the disease, and the accumulation of amyloid A β in the brain, the increased number and the misfolding of Aβ are generally thought to be directly related to Alzheimer's disease. Studies have shown that A β can be mistakenly folded into fibrous aggregates rich in beta-folded structures, which have neurotoxicity. The aim of this study is to express β
-amyloid protein by prokaryotic expression of Escherichia coli. The experimental method was based on the existing Aβ40 and Aβ42 gene templates and primers. The Aβ42 gene fragment was amplified by PCR and cloned into the vector plasmid pET-28a. The recombinant plasmid pET-28a-Aβ42 was transformed into Escherichia coli DH5 α , and the positive clones were induced by IPTG. After restriction enzyme digestion and DNA sequencing, the recombinant expression vector was successfully constructed. The expression of [beta]-amyloid protein was shown by SDS-PAGE electrophoresis. The A β 40 and A β 42 genes were successfully cloned and the expression was stable in Escherichia coli. This study provides material and technical support for the further production of recombinant beta-amyloid protein and Alzheimer's disease, and supports the study of the structural function and dynamic process of Aβ.
Keywords Alzheimer's disease Beta-amyloid protein gene expression clone
目 录
1 引言 1
1.1 分子生物学基础 1
1.2 β淀粉样蛋白（Aβ） 1
1.2.1 Aβ的结构和形成 4
1.2.2 淀粉样蛋白的级联假说 4
1.3 多肽固相合成法和点击化学 5
1.3.1 多肽固相合成法 5
1.3.2 点击化学简介 9
1.4 阿尔茨海默病的免疫治疗 阿尔兹海默病模型多肽的化学合成与生物表达:http://www.chuibin.com/yixue/lunwen_205385.html