由特异基因扩增产物制备丙型肝炎病毒cDNA探针  

Preparation of Hepatitis C Virus cDNA Probe from a Sequence Amplified by Polymerase Chain Reaction(PCR)

作　　者：吕凌[1] 姚集鲁[1] 彭文伟 

机构地区：[1]广州中山医科大学附属第二医院传染病科

出　　处：《中华实验和临床病毒学杂志》1995年第3期217-220,共4页Chinese Journal of Experimental and Clinical Virology

摘　　要：应用逆转录聚合酶链反应技术自丙型肝炎病毒（ＨＣＶ）感染血浆中扩增ＨＣＶ－ｃＤＮＡ，经寡聚核苷酸探针杂交鉴定后以低熔点琼脂糖凝胶电泳分离、微柱亲和层析纯化，用于缺口平移制备（３２）Ｐ标记的ＨＣＶ基因探针。这种探针用于斑点杂交和Ｓｏｕｔｈｅｒｎ杂交检测非甲非乙型肝炎（ＮＡＮＢＨ）血浆的ＨＣＶ基因扩增产物，能鉴定ＨＣＶ序列的特异性，且分别达到０．１ｐｇ和１ｐｇ的敏感性。这种方法基因来源方便，纯化制备简单，且不失克隆ｃＤＮＡ优点，因此有推广应用价值。A sequence was amplified from hepatitis C virus (HCV) genome by polmerase chain reaction (PCR) from plasma of HCV-infected patients with its specificity determined by an oligonucleotide derived from the Taiwan Residents strain HCV. After electrophoretic separation in low melting point agarose gel and elution from a minicolumn of affinity chromotography,it was used in nick translation to prepare (32)P-labelled HCV-cDNA probe. By applying in spot-blot and Southern-blot hybridization, the probe can specificically detect its homologous sequence from another PCR with a reactive sensitivity of 0.1 pg and 1 pg. The described method is a practical skill in that it offers a convenient supply of HCV sequence which shares the same characteristics with the cloned HCV-cDNAs and from which HCV probe can simply be generated that can be applied in the detection of PCR products.

关 键 词：丙型肝炎病毒 DNA探针 基因扩增 制备 

分 类 号：R373.21[医药卫生—病原生物学] R392－33[医药卫生—基础医学]