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带GFP标签LMO4质粒的构建及转染表达

时间:2020-01-16 19:09来源:毕业论文
为了对目的基因LMO4进行示踪,通过基因重组技术,构建能够表达绿色荧光蛋白基因(GFP)的大肠杆菌菌株。方法:构建LMO4-GFP质粒,以大肠杆菌D5Hα菌株作为感受态进行转化培养

摘要最新的研究表明LMO4是耳蜗感觉器官形成的负调控因子。基于LMO家族蛋白与LIM-HD竞争结合LDB蛋白的特性, LMO4可能对某种耳蜗产生关键因子的功能起到调控作用。这就提示着LMO4极有可能在抑制耳蜗腹侧上皮细胞中产生prosensory domain的能力。目的:为了对目的基因LMO4进行示踪,通过基因重组技术,构建能够表达绿色荧光蛋白基因(GFP)的大肠杆菌菌株。方法:构建LMO4-GFP质粒,以大肠杆菌D5Hα菌株作为感受态进行转化培养,观察绿色荧光蛋白的表达情况。结果:在荧光显微镜下检测到绿色荧光表达,成功构建LMO4-GFP质粒。44085

Abstract The latest research shows that LMO4 is a negative regulator of the formation of cochlear sensory organ. Based on the ability of LMO-family proteins to compete against LIM-homeodomain factors in binding to LDB proteins, it is conceivable that LMO4 could regulate some key factors to produce cochlea. It is suggests that LMO4 in the ventral cochlear epithelium could inhibit the ability to generate prosensory domain. Objective: For the purposes of gene LMO4 tracer, through recombinant DNA technology, constructed to express green fluorescent protein (GFP) strain of E. coli.  METHODS: LMO4-GFP plasmid was constructed, and the E. coli strains were transformed culture D5Hα as competent to observe the expression of green fluorescent protein. Results: green fluorescent expression was detected by fluorescence microscopy, and LMO4-GFP plasmid was constructed successfully.

毕业论文关键词:LMO4; 内耳; GFP; 质粒构建; 荧光检测

Keyword: LMO4; Inner ear; GFP; Plasmid Construction; Fluorescence detection

目录

1. LMO4 5

1.1. LMO4的传统功能研究 5

1.2. LMO4在内耳形态发生中的作用 5

1.3. LMO4的最新研究进展及展望 6

2. GFP标签 7

2.1. GFP标签的发光机制 7

3. 实验内容与设计 8

3.1. 实验目的 8

3.2. 实验内容 8

3.3. 实验流程图 8

4. 实验方法 9

4.1. 提取RNA及反转录 9

4.1.1. 提取RNA 9

4.1.2. RNA反转录步骤 : 9

4.2. 目的基因片段PCR 10

4.3. PCR产物与质粒的酶切与连接 11

4.4. 质粒的大提及纯化 13

4.5. 细胞转染 14

4.6. 荧光检测 14

5. 实验总结与讨论 14

5.1. 结论 带GFP标签LMO4质粒的构建及转染表达:http://www.lwfree.com/shengwu/lunwen_45255.html

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