艾滋病痴呆综合征患者HIV-1 nef基因多样性分析

摘要

Objective To study the genetic diversity of HIV-1 nef genes from a patient with AIDS dementia complex (ADC),so as to research the amino acid variability and the pathogenesis of ADC.Methods The nef gene was amplified with PCR from genomic DNA which was extracted from spleen and different brain tissues(basal ganglia,frontal gray matter,meninges,temporal lobe)of a patient who died of ADC.PCR products were cloned into the pMD19-T vector,after transformation and selection by ampicillin and blue/white spotting.Five of positive clones were sequenced and confirmed with BLAST.HIV-1 nef sequences were processed with BioEdit and MEGA4 to do Neighbor-Joining tree,p-Distances,and values of ds/dn.Results The samples were all identified as HIV-1 B and genetic variation exists in HIV-1 nef gene isolated from different tissues compared with HXB2.In addition,part of the changes were different between periphery and brain.Conclusion Variations exist in the HIV-1 nef gene extracted from the ADC patient and the variations from peripheral and central nerve tissues were different,these variations may change the function of Nef,and it needs more research.%目的 研究艾滋病痴呆综合征(ADC)患者体内HIV-1 nef基因变异规律,为ADC发病机制的研究提供依据.方法 提取1例ADC病例尸检标本的外周(脾)和中枢神经系统(基底核、额叶灰质、脑膜、颞叶)共5个部位组织中的基因组DNA,PCR扩增HIV-1 nef基因,与pMD19-T克隆载体连接后,蓝白斑筛选挑取阳性菌落测序,序列经BLAST分析后,每个部位取5个序列,利用Bioedit、MEGA4等生物学软件进行基因距离、系统进化树以及同义/非同义替换值(ds/dn)等分析.结果 该ADC患者感染的是HIV-1 B亚型,与标准序列HXB2比较,该病例的HIV-1 nef基因存在变异,且外周组织与中枢神经系统各组织间以及中枢神经组织不同部位的基因变异不同,相同组织来源的nef序列基因距离较小,外周和中枢的nef基因距离差异较大,该病例H IV-1 nef基因在外周和中枢神经系统变异时均受到负向选择压力.结论 ADC患者体内HIV-1 nef基因存在多样性,且不同组织部位的基因变异不同,其导致的Nef蛋白某些氨基酸位点的改变对其生物学活性的影响有待进一步研究.