CAJ | 학술논문

目的克隆艾滋病痴呆综合征(ADC)患者体内不同部位的HIV-1B gp120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-1 gp120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/pcDNA3.1(+),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-IB gp120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-1B gp120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-1B gp120基因均可在U87细胞中表达,为进一步研究HIV-1B gp120包膜蛋白的神经毒性及其作用机制提供条件.
목적 극륭애자병치태종합정(ADC)환자체내불동부위적HIV-1B gp120기인,병재인신경효질류세포U87중표체.방법 분별이일례ADC환자시검표본적외주(림파결)화중추(맥락총、대뇌침협백질)래원적기인조DNA위모판,PCR확증HIV-1 gp120기인,서렬측정후,장목적기인삽입표체재체pcDNA3.1(+),구건중조표체재체gp120/pcDNA3.1(+),장소구건적중조표체재체용지질체법전염인신경효질류U87세포,간접면역형광법측정HIV-IB gp120적표체정황.결과 극륭료ADC환자체내외주림파결、맥락총、대뇌침협백질3개부위적HIV-1B gp120기인,소구건적표체질립gp120/pcDNA3.1(+)전염U87세포후가표체목적단백.결론 ADC환자불동부위래원적HIV-1B gp120기인균가재U87세포중표체,위진일보연구HIV-1B gp120포막단백적신경독성급기작용궤제제공조건.
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,HIVIB gp120 was inserted into pcDNA3.1 ( + ) and recombinant expressing vector gp120/pcDNA3.1 ( + ) was constructed succeffuly confirming with sequencing. Then expressing vector was transfected into eukaryotic cells U87 using liposome transfection and expression of HIV-1B gp120 gene was assayed with indirect immunofluorescence.Results HIV-I B gp120 genes isolated from peripheral lymphnodes,choroid plexus and occipital white matter of the ADC patient were successfully cloned and recombinant expressing vector gp120/pcDNA3.i ( + ) could express envelope glycoprotein HIV-1B gp120 in U87 cells.Conclusion All the HIV-1B gp120 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 gp120 protein.