含密码子优化型HIV-1 gp120基因重组腺病毒的构建及其免疫效果研究  

Construction and immune potency of recombinant adenovirus containing codon-modified HIV-1 gp120

作　　者：冯霞[1] 余双庆[1] 陈国敏[1] 左建民[1] 周玲[1] 曾毅[1] 

机构地区：[1]中国疾病预防控制中心病毒病预防控制所肿瘤病毒与艾滋病病毒研究室,北京100052

出　　处：《中华实验和临床病毒学杂志》2004年第2期113-117,共5页Chinese Journal of Experimental and Clinical Virology

基　　金：中国综合性艾滋病研究项目 (CIPRA) ;国家 863计划项目 (2 0 0 1AA2 15 0 3 1)

摘　　要：目的　构建能表达野生型和密码子优化型人免疫缺陷病毒Ⅰ型 (HIV 1)B亚型中国流行株gp12 0基因的非复制型腺病毒。方法　按哺乳动物细胞偏好的密码子对HIV 1B亚型中国流行株Chgp4 2的gp12 0基因进行优化 ,合成优化基因。将野生型和密码子优化的gp12 0基因插入穿梭质粒 ,再与腺病毒骨架质粒pAdEasy 1共转化E coliBJ5 183,获得重组子 ,转染 2 93细胞后获得重组病毒。分别以两种重组腺病毒疫苗免疫小鼠 ,ELISA检测小鼠血清中的特异性抗体 ,乳酸脱氢酶法检测小鼠细胞毒性T淋巴细胞 (CTL)反应。结果　获得两株重组腺病毒rAd wt gp12 0和rAd mod gp12 0 ,能正确表达Gp12 0。rAd mod gp12 0比rAd wt gp12 0蛋白表达水平明显提高。重组腺病毒免疫小鼠后能产生HIV 1特异性的抗体及CTL反应 ,rAd mod gp12 0组明显优于rAd wt gp12 0组。结论　成功构建了表达野生型和密码子优化的HIV 1gp12 0基因的重组腺病毒 ,能诱导HIV 1特异性体液和细胞免疫反应。Objective To construct replication-deficient recombinant adenovirus expressing wild and codon-modified HIV-1 gp120. Methods The viral codons were changed to the codon usage of highly expressed mammal gene,the resulting modified gp120 gene was synthesized. The wild and modified gp120 genes were cloned into shuttle vector pShuttle-CMV respectively,and then the constructed plasmids containing gp120 gene was cotransformed with the backbone vector pADeasy-1 into E.coli BJ5183. Transfection of the recombinant AdEasy plasmid into 293 cells was performed to obtain recombinant adenoviruses. The mice were immunized with the recombinant adenoviruses. Their immunogenicity was evaluated by testing antibody and CTL levels of immunized mice. Results Two strains of recombinant adenovirus expressing wild and codon-modified HIV-1 gp120 were obtained. The protein expressing level of the recombinant adenoviruses containing modified genes was much higher than that containing wild genes. The mice immunized with recombinant adenoviruses elicited HIV-1 specific antibody and CTL response. The rAd-mod.gp120 group was better than the rAd-wt.gp120 group. Conlusion Replication-deficient recombinant adenovirus expressing HIV-1 gp120 can elicit HIV-1 specific humoral and cellular response,the condon-modified recombinant virus was more efficient than the native.

关 键 词：HIV-1 GP120 重组腺病毒 免疫效果 人免疫缺陷病毒Ⅰ型 密码子优化型 野生型 基因优化 

分 类 号：R392[医药卫生—免疫学]