Alu-反向PCR检测HIV-1整合位点方法的建立  

Detecting HIV-1 integration site by Alu-Inverse PCR

作　　者：田亚坤[1] 李侗曾[1] 焦艳梅[1] 李娟[1] 画伟[1] 吴昊[1] 

机构地区：[1]首都医科大学附属北京佑安医院感染科,100069

出　　处：《北京医学》2010年第12期965-968,共4页Beijing Medical Journal

基　　金：国家科技重大专项(2008ZX10001-006);北京市科委重大项目(D09060030405912)资助项目

摘　　要：目的建立Alu-反向PCR检测整合型HIV-1DNA的实验方法。方法收集18例HIV-1感染者及12例健康者的抗凝血标本,根据整合型HIV-1DNA的特点设计引物,第一轮PCR5’端引物来自于人类保守的Alu序列,3’端引物来自于HIV-1gag序列,扩增片段包含了整合位点上游的人类基因组DNA序列和整合的HIV-1DNA序列。经PstⅠ酶切后进行自身片段环化,以此为模板设计针对HIV-1LTR和gag保守区域引物,进行巢式PCR。结果 16例HIV-1感染者成功扩增出目的片段,12例健康者均为阴性。结论本实验建立的检测整合型HIV-1的方法为进一步研究HIV病毒储藏库机制提供了参考。Objective To establish a study method that measure HIV integration DNA.Methods The blood samples were collected from 18 HIV-1 infected patients and 12 healthy people,A pair of primers were designed according to characteristics of human and HIV-1 genome.The first PCR was carried out by the nested 5＇ primers from conserved sequences of human Alu and 3＇ primer from HIV-1 gag sequences.The product of the first PCR was representative of both cellular DNA upstream of the integration site and seguneces of integrated HIV-1.The product was firstly digested with PstⅠ,and then intramolecular ligeted and nested amplified.Results Integrated HIV-1 sequence was detected in each of 16 cases of HIV/AIDS and non in healthy peoples.Conclusion The study method provides a support to research viral reservoir.

关 键 词：HIV-1 整合型HIV Alu序列 反向PCR 

分 类 号：R512.91[医药卫生—内科学]