艾滋病痴呆综合征患者HIV-1B gp120基因的克隆及真核表达  

Cloning and eukaryotic expression of HIV-1B gp120 genes from a patient with AIDS dementia complex

作　　者：赵丽[1] 闫玉芬[2] 李静[3] 蒲双双[4] 王志玉[1] 温红玲[1] 宋艳艳[1] 许洪芝[1] 

机构地区：[1]山东大学公共卫生学院微生物检验研究所,济南250012 [2]青岛大学医学院附属医院黄岛分院预防保健科 [3]青岛市市立医院检验科 [4]山东中医药大学第一附属医院检验科

出　　处：《中华实验和临床病毒学杂志》2012年第2期93-95,共3页Chinese Journal of Experimental and Clinical Virology

基　　金：山东省科技发展项目资助（2007GG30002003）;山东大学公共卫生学院青年人才创新基金资助

摘　　要：目的克隆艾滋病痴呆综合征（ADC）患者体内不同部位的HIV-1Bgpl20基因，并在人神经胶质瘤细胞U87中表达。方法分别以一例ADC患者尸检标本的外周（淋巴结）和中枢（脉络丛、大脑枕叶白质）来源的基因组DNA为模板，PCR扩增HIV-1gp120基因，序列测定后，将目的基因插入表达载体pcDNA3．1（＋），构建重组表达载体gp120／peDNA3．1（＋），将所构建的重组表达载体用脂质体法转染人神经胶质瘤U87细胞，间接免疫荧光法测定HIV-1Bgp120的表达情况。结果克隆了ADC患者体内外周淋巴结、脉络丛、大脑枕叶白质3个部位的HIV-1Bgp120基因，所构建的表达质粒gp120／pcDNA3．1（＋）转染U87细胞后可表达目的蛋白。结论ADC患者不同部位来源的HIV-1Bgp120基因均可在U87细胞中表达，为进一步研究HIV-1Bgp120包膜蛋白的神经毒性及其作用机制提供条件。Objective To clone and express the HIV-1B gp120 genes isolated at different organizations from a patient died of AIDS dementia complex （ADC） in eukaryotic cells. Methods Using the genomic DNA isolated from peripheral lymphnodes, choroid plexus and occipital white matter from a patient died of ADC as the template, HIV-1B gp120 gene was amplified with PCR. After sequenced, HIV- 1B gpl20 was inserted into pcDNA3. 1 （ ＋ ） and recombinant expressing vector gpl20/pcDNA3.1 （ ＋ ） was constructed succeffuly confirming with sequencing. Then expressing vector was transfected into eukaryotic cells U87 using liposome transfection and expression of HIV-IB gpl20 gene was assayed with indirect immunofluorescence. Results HIV-IB gpl20 genes isolated from peripheral lymphnodes, choroid plexus and occipital white matter of the ADC patient were successfully cloned and recombinant expressing vector gpl20/pcDNA3. 1 （ ＋ ） could express envelope glycoprotein HIV-IB gpl20 in U87 cells. Conclusion All the HIV-IB gpl20 gene isolated at the different organizations of the same ADC patient could express in U87 cells, which may supply a valuable basis for studying the neurotoxicity and neurotoxic mechanism of HIV-1 gpl20 protein.

关 键 词：获得性免疫缺陷综合征 HIV-1 克隆 分子 表达的序列标记 

分 类 号：R512.91[医药卫生—内科学]