荧光标记重组乙型肝炎病毒载体的构建

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1、授予单位代码10089学号或申请号20122426HebeiMedicalUniversity硕士学位论文科学学位荧光标记重组乙型肝炎病毒载体的构建研究生：张银阁导师：孙殿兴教授专业：内科学二级学院：白求恩国际和平医院2015年3月河北医科大学学位论文使用授权及知识产权归属承诺本学位论文在导师（或指导小组）的指导下，由本人独立完成。本学位论文研究所获得的研究成果，其知识产权归河北医科大学所有。河北医科大学有权对本学位论文进行交流、公幵和使用。凡发表与学位论文主要内容相关的论文，第一署名为单位河北医科大学，试验材料、原始数据、申报

2、的专利等知识产权均归河北医科大学所有。否则，承担相应法律责任。研究生签名：丨^if师签章^i/nz级第囊翻导盖章1“in河北医科大学研究生学位论文独创性声明本论文是在导师指导下进行的研究工作及取得的研究成果，除了文中特别加以标注和致谢等内容外，文中不包含其他人已经发表或撰写的研究成果，指导教师对此进行了审定。本论文由本人独立撰写，文责自负。研究生签名：丨知{如今导师签章：^、H乂年3月^曰目录中文摘要·································································

3、··············1英文摘要···············································································5英文缩写·············································································10研究论文荧光标记重组乙型肝炎病毒载体的构建前言······························································

4、···············11材料与方法····································································12结果·············································································24附图·············································································29附表···········

5、··································································40讨论·············································································41结论·············································································45参考文献·································

6、······································45综述绿色荧光蛋白标记技术及其应用······································49致谢···················································································57个人简历·············································································58中

7、文摘要荧光标记重组乙型肝炎病毒载体的构建摘要目的：把乙型肝炎病毒（HepatitisBvirus,HBV）改造为携带外源基因的复制型载体，可视化的观察HBV的表达与复制。方法：1利用PCR和分子克隆技术构建表达切开绿色荧光蛋白（greenfluorescentprotein,GFP）1-10片段的质粒psuCMV-GFP1-10；构建表达GFP11片段的载体质粒：pCH-GFP11-Core,pCH-GFP11-BsdR和pCH-GFP11-S2。2质粒psuCMV-GFP1-10分别与表达GFP11的载体质粒pCH-GFP11

8、-Core,pCH-GFP11-BsdR和pCH-GFP11-S2共转染肝癌细胞系HepG2细胞，观察GFP表达的强度和特点。3载体质粒pCH-GFP11-Core,pCH-GFP11-BsdR和pCH-GFP11-S2分别转染HepG2细胞，转染后第五天提取细