摘要
作者设计了一对DNA引物,用于扩增1型人免疫缺陷病毒(HIV—1)gag基因的一段保守序列。应用聚合酶链反应(PCR)技术,采用含有ARV-2gag基因的亚克隆质粒pAG423作模板,可以使靶基因片段数目增加1×10~7倍以上,经电泳染色后,扩增产物清晰可辨。实验结果表明,这种方法的敏感性极高,足以检测到6.5个拷贝的HIV-1基因。用特异性DNA探针打点杂交证实扩增产物为特异性gag基因片段。实验还表明,经过30～35次PCR循环可以得到最佳扩增效果,而且应用PCR和打点杂交方法还可以对HIV-1基因进行定量分析。因而PCR是一种可以取代病毒分离的常规测定HIV-1感染的简便方法。
A conserved sequence of human immunodeficiency virus type 1 (HTV - 1) gag gsne was amplified using a selective DNA amplification technique, pdymerase chain reaction (PCR). Using pAG423 that contains gag gene of ARV - 2 as template, by PCR technique, the target sequence of HTV 1 gene could be amplified over 10' times and was identified simply by electro phoresis and ethydium bromide staining. It was found for the first time that the sensitivity of PCR was high enough to detect a few copies of HTV - 1 gene directly. The product of amplification was identified by dot blot hybridization with a specific probe. Usually the best amplification was obtained by 30 to 35 cycles of PCR. If unsaturated rounds of amplification was used, quantitative analysis of HIV - 1 provirus was proved possible . The results suggest that PCR is a simple method instead of virus isolation for the determination of HTV -1 infection.
出处
《第四军医大学学报》
1990年第6期405-408,共4页
Journal of the Fourth Military Medical University
关键词
聚合酶链反应
HIV-1
核酸杂交
pdymerasc chain reaction
human immunodeficiency virus
plasmid
nudcic acid hybridization
