采用TaqMan MGB探针定量检测HCV 1b基因型C区氨基酸70变异  

Establishment of a quantification method for the amino acid 70 substitution in the core region of hepatitis C virus genotype 1b

作　　者：胡中杰[1] 刘颖[1] 仇丽霞[1] 范作鹏[1] 聂巍[1] 梁珊[1] 

机构地区：[1]首都医科大学附属北京佑安医院丙肝与中毒性肝病科,100069

出　　处：《中华实验和临床病毒学杂志》2014年第4期305-308,共4页Chinese Journal of Experimental and Clinical Virology

基　　金：北京市优秀人才培养资助（No.2010D003034000009）;首都医科大学基础·临床科研合作基金（NO,1000172053-11JL61）;中国初级保健基金会佑安肝病艾滋病摹金（NO.BJYAH-2011-073）

摘　　要：目的 建立一种定量检测HCV 1b基因型C区氨基酸70位点主要野生株（Arg-CGG,70w）和变异株（GIn-CAG,70M）的方法.方法 设计一对分别针对70 W和70 M的TaqMan MGB探针,建立一种实时逆转录聚合酶链反应定量检测方法.验证方法的敏感性、特异性和准确性.结果 所设计的引物和探针具有良好的特异性,敏感性可达5拷贝/反应.进一步的克隆测序证实了新方法的可靠性.结论 建立了一套HCV 1b亚型C区氨基酸70变异的定量检测系统,具有较高的敏感性、特异性和准确性,为研究70 W和70 M在抗病毒治疗中的动态变化提供了技术手段.Objective To develop a rapid and sensitive method to quantify the substitution of amino acid （aa） 70 in hepatitis C virus （HCV） genotype 1 b core region.Methods Specific primers and TaqManminor groove binder probes were designed to establish a real time reverse transcription-polymerase chain reaction for aa70 substitution.The specificity,sensitivity and accuracy of the system were tested.Results The probes could clearly distinguish 70W and 70M.The detection limit for each system was 5 copies/ reaction.Further tests using cloning sequencing confirmed the reliability of the detection system.Conclusions A novel system was developed and provides an effective,sensitive,and accurate approach to quantify the wild type and mutant strains of HCV 1b core region as70.

关 键 词：肝炎病毒属 逆转录聚合酶链反应 

分 类 号：R512.63[医药卫生—内科学]