Feng Xia, Yu Shuang-qing, Chen Guo-min, Zuo Jian-min, Zhou Ling, Zeng Yi
Department of Oncogenic Virus and HIV, Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 100052, China.
Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi. 2004 Jun;18(2):113-7.
To construct replication-deficient recombinant adenovirus expressing wild and codon-modified HIV-1 gp120.
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.
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.
Replication-deficient recombinant adenovirus expressing HIV-1 gp120 can elicit HIV-1 specific humoral and cellular response, the codon-modified recombinant virus was more efficient than the native.
构建表达野生型和密码子优化型HIV-1 gp120的复制缺陷型重组腺病毒。
将病毒密码子替换为高表达哺乳动物基因的密码子使用方式，合成得到修饰后的gp120基因。将野生型和修饰后的gp120基因分别克隆至穿梭载体pShuttle-CMV，随后将构建好的含gp120基因的质粒与骨架载体pADeasy-1共转化至大肠杆菌BJ5183。将重组AdEasy质粒转染至293细胞以获得重组腺病毒。用重组腺病毒免疫小鼠。通过检测免疫小鼠的抗体和CTL水平评估其免疫原性。
获得了两株表达野生型和密码子优化型HIV-1 gp120的重组腺病毒。含修饰基因的重组腺病毒的蛋白表达水平远高于含野生型基因的重组腺病毒。用重组腺病毒免疫的小鼠引发了HIV-1特异性抗体和CTL反应。rAd-mod gp120组优于rAd-wt gp120组。
表达HIV-1 gp120的复制缺陷型重组腺病毒可引发HIV-1特异性体液和细胞反应，密码子优化型重组病毒比天然型更有效。
