丙型肝炎病毒结构基因转基因小鼠模型的建立  

Establish a transgenic mice model harboring structural genes of hepatitis C virus

作　　者：任进余[1] 成国祥 孔晓飞[1] 陈建泉 周汝江 陆志檬[1] 

机构地区：[1]上海第二医科大学附属瑞金医院临床病毒研究室,200025 [2]上海转基因研究中心

出　　处：《中华肝脏病杂志》2005年第7期501-504,共4页Chinese Journal of Hepatology

基　　金：上海市科委重点资助项目(984119007)

摘　　要：目的建立丙型肝炎病毒(HCV)转基因小鼠模型,探讨丙型肝炎发病机制,研究HCV结构蛋白编码基因的功能与致病作用。方法采用长片段聚合酶链反应技术扩增HCV结构基因,构建稳定表达与诱导表达两种真核细胞表达重组体,经体外细胞转染后,将具有表达功能的线性化表达元件采用显微注射法,直接导入1736枚小鼠受精卵,先后移入69只假孕母鼠输卵管。结果妊娠受体25只,产仔105只,成活57只。注射卵成仔率仅3.2 8%。经聚合酶链反应扩增、测序、酶切及Southern杂交鉴定,获阳性首建鼠26只,注射卵首建鼠成功率仅1.50%。其中稳定表达型10只,诱导表达型16只。经与正常鼠交配,获F1阳性杂合鼠分别38只、20只。一个纯合子品系。结论研究建立了稳定表达与诱导表达两类HCV 结构基因转基因小鼠,为进一步研究HCV致病机制,HCV基因功能奠定了基础。Objective To establish an animal model of HCV transgenic mice to elucidate the pathogenesis of hepatitis C virus infection and function of the viral structural proteins. Methods Structural gene of HCV were amplified and recombined into eukaryotic expression vectors, pcDNA4HisMax and pMT/BiP/V5-His A, after their expressive activity was confirmed to detect the structural protein in the transfected COS7 and S2 cells by Western blot. The fertilized expression element, which contained CMV or pMT promoter, structural gene of HCV and polyadenylation signal sequence, was microinjected into 1736 C57BL/6 mouse fertilized ova. The ova were then replanted into the oviducts of 69 pseudopregnant recipient mice. Results Twenty-five recipient mice were impregnanted and later produced 105 newborns; 49 of them died from unknown causes and 57 survived. After the specific HCV structural genes were identified by PCR and Southern blot hybridization, 26 founders were obtained; among them 10 were stable expression mice and 16 were the inducible ones. The rate of founders developed from implanted embryos was only 1.50%. Through hybridization with normal mice, 58 hybrid mice have been obtained at present. Conclusion Two kinds of different transgenic mice of HCV were developed; one is of stable expression, and the other is inducible. This transgenic mice model may create an opportunity for studying the function of the structural gene of HCV and elucidate its pathogenicity.

关 键 词：丙型肝炎 病毒结构基因 转基因 小鼠模型 聚合酶链反应 HCV基因 

分 类 号：R-332[医药卫生]