乙型肝炎病毒基因（S、X）載體構建表達及功能研究
發(fā)布時間：2021-09-09 19:58
目的分別構建乙型肝炎病毒（Hepatitis B Virus，HBV）S基因（包括Pre-S1，Pre-S2和S）和X基因的帶FLAG標簽的真核表達載體，并進行有效表達，進而通過高通量蛋白質(zhì)技術獲取與X蛋白相互作用的蛋白質(zhì)復合物，以期為進一步研究乙型肝炎病毒外膜蛋白及X蛋白的功能打下基礎。方法以質(zhì)粒pcDNA3.1-HBV為模板，通過PCR技術擴增HBV基因組Large surface antigen（LS）、Middle surface antigen（MS）、Smallsurface antigen（SS）和X基因片段；將PCR克隆獲得的目的基因片段分別插入真核表達載體pCMV-tag2B，構建重組質(zhì)粒pCMV-tag2B-LS、 pCMV-tag2B-MS、 pCMV-tag2B-SS和pCMV-tag2B-X，并送測序鑒定。將鑒定正確后的目的基因表達載體分別轉染293FT細胞，Western blot檢測其表達。繼而應用FLAG親和層析柱分離HBX復合體，iTRAQ標記后用串聯(lián)質(zhì)譜對其進行鑒定。選擇三次實驗中只在HBX復合體中鑒定的蛋白質(zhì)作為進一步功能分析的侯選蛋白。通過免疫共...
【文章來源】：重慶醫(yī)科大學重慶市
【文章頁數(shù)】：50 頁
【學位級別】：碩士
【部分圖文】：
凝膠電泳分離LS、MS、SS、X的PCR產(chǎn)物
圖 1.1 凝膠電泳分離 LS、MS、SS、X 的PCR 產(chǎn)物Fig 1.1 PCR product of LS、MS、SS and Xseperated by Agar Gel electrophoresis圖 1.2 pCMV-tag2b-LS, pCMV-tag2b-MS,pCMV-tag2b-SS 雙酶切后Fig 1.2 pCMV-tag2b-LS, pCMV-tag2b-MS,pCMV-tag2b-SS after digested by restrictionenzyme
圖 1.1 凝膠電泳分離 LS、MS、SS、X 的PCR 產(chǎn)物Fig 1.1 PCR product of LS、MS、SS and Xseperated by Agar Gel electrophoresis圖 1.2 pCMV-tag2b-LS, pCMV-tag2b-MS,pCMV-tag2b-SS 雙酶切后Fig 1.2 pCMV-tag2b-LS, pCMV-tag2b-MS,pCMV-tag2b-SS after digested by restrictionenzyme
【參考文獻】：
期刊論文
[1]Adenovirus-expressed preS2 antibody inhibits hepatitis B virus infection and hepatic carcinogenesis[J]. Qian Zhang,Department of Infectious Diseases,Changhai Hospital,Second Military Medical University,Shanghai 200433,China Zhi-Qing Li,Hu Liu,Jia-He Yang,Department of Hepatic Surgery,Eastern Hepatobiliary Surgery Hospital,Second Military Medical University,Shanghai 200438,China. World Journal of Gastroenterology. 2012(04)
[2]Hepatitis B virus X protein promotes proliferation and upregulates TGF-β1 and CTGF in human hepatic stellate cell line,LX-2[J]. Guang-Hui Guo,De-Ming Tan,Ping-An Zhu and Fei Liu Department of Infectious Diseases,Xiangya Hospital,Central South University,Changsha 410008,China. Hepatobiliary & Pancreatic Diseases International. 2009(01)
本文編號：3392694
【文章來源】：重慶醫(yī)科大學重慶市
【文章頁數(shù)】：50 頁
【學位級別】：碩士
【部分圖文】：
凝膠電泳分離LS、MS、SS、X的PCR產(chǎn)物
圖 1.1 凝膠電泳分離 LS、MS、SS、X 的PCR 產(chǎn)物Fig 1.1 PCR product of LS、MS、SS and Xseperated by Agar Gel electrophoresis圖 1.2 pCMV-tag2b-LS, pCMV-tag2b-MS,pCMV-tag2b-SS 雙酶切后Fig 1.2 pCMV-tag2b-LS, pCMV-tag2b-MS,pCMV-tag2b-SS after digested by restrictionenzyme
圖 1.1 凝膠電泳分離 LS、MS、SS、X 的PCR 產(chǎn)物Fig 1.1 PCR product of LS、MS、SS and Xseperated by Agar Gel electrophoresis圖 1.2 pCMV-tag2b-LS, pCMV-tag2b-MS,pCMV-tag2b-SS 雙酶切后Fig 1.2 pCMV-tag2b-LS, pCMV-tag2b-MS,pCMV-tag2b-SS after digested by restrictionenzyme
【參考文獻】：
期刊論文
[1]Adenovirus-expressed preS2 antibody inhibits hepatitis B virus infection and hepatic carcinogenesis[J]. Qian Zhang,Department of Infectious Diseases,Changhai Hospital,Second Military Medical University,Shanghai 200433,China Zhi-Qing Li,Hu Liu,Jia-He Yang,Department of Hepatic Surgery,Eastern Hepatobiliary Surgery Hospital,Second Military Medical University,Shanghai 200438,China. World Journal of Gastroenterology. 2012(04)
[2]Hepatitis B virus X protein promotes proliferation and upregulates TGF-β1 and CTGF in human hepatic stellate cell line,LX-2[J]. Guang-Hui Guo,De-Ming Tan,Ping-An Zhu and Fei Liu Department of Infectious Diseases,Xiangya Hospital,Central South University,Changsha 410008,China. Hepatobiliary & Pancreatic Diseases International. 2009(01)
本文編號：3392694
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