摘要
将HIV-2嵌合结构基因gag-gp105插入到转移载体pUTA2复合启动子ATI-P7.5×20下游,构建出鸡痘病毒重组转移质粒pA-gg;经转染和BrdU加压筛选,以基因组PCR和Western blot法鉴定重组病毒;将获得的重组病毒大量制备并纯化后,肌肉注射免疫BALB/c小鼠,ELISA法检测小鼠血清HIV-2抗体,流式细胞仪测定CD4^＋、CD8^＋T淋巴细胞亚类数量,乳酸脱氢酶（LDH）释放法检测脾特异性CTL杀伤活性.结果表明,成功筛选出一株可稳定表达HIV-2嵌合蛋白Gag-gp105的重组鸡痘病毒rFPV-gg;该重组病毒免疫组小鼠的血清出现HIV-2抗体,脾T细胞亚类数量增加,并产生针对HIV-2靶细胞的特异性CTL杀伤活性.本研究提示,HIV-2 gag-gp105重组病毒能诱导小鼠产生特异性细胞和体液免疫应答.
A fowlpox virus （FPV） transferring vector pUTA2-gag-gp105 was constructed by inserting HIV-2 gag-gpl05 chimeric gene to the downstream of a synthetic complex promoter ATI-p7.5× 20 of vector pUTA2. Transfection was then carried out, and recombinant FPV （rFPV） was screened by 5＇-bromo-deoxym＇idine （BrdU）, genome PCR and Western blot detection. Hereditary stability of the rFPV was detected. BALB/c mice were immunized with rFPV by muscular injection. Anti-HIV-2 antibody, CD^4＋ and CD^8＋ T-cell count and specific target-killing activity of spleen CTL in immunized mice were analyzed by ELISA, FACS and LDH release assay, respectively. The results showed that a recombinant virus rFPV- gg which could stablely express HIV-2 chimeric protein gag-gpl05 was obtained. The HIV-2 specific antibody was detect- ed from the immunized BALB/c mice. The numbers of CD^4＋ and CD^8＋ subgroup of spleen T lymphocyte were higher in immunization group than those in controls. HIV-2-Specific target-killing activity of spleen CTL was observed in immunized mice. It was concluded that the rFPV-gg may elicit specific celluar and humoral immune reactions in mice.
出处
《高技术通讯》
CAS
CSCD
北大核心
2006年第7期730-734,共5页
Chinese High Technology Letters
基金
863计划（2003AA219051）、吉林省科技发展计划（20030550-1）资助项目.
