引物标记与掺入标记在HBV基因多态性芯片检测中应用的研究  

Influence of labeled primer and labeled dUTP assays on the signal intensity of the chip for the detection of HBV gene polymorphism

作　　者：马达 王万相 郭乃洲 蒋玲 王惠民[2] 张冬雷[2] 赵建龙[3] 孙悦[3] 

机构地区：[1]江苏省盐城市第一人民医院,盐城224001 [2]南通医学院附属医院 [3]中国科学院上海冶金研究所

出　　处：《中华实验和临床病毒学杂志》2002年第2期166-167,共2页Chinese Journal of Experimental and Clinical Virology

摘　　要：目的　研究引物标记与掺入标记在乙型肝炎病毒 (HBV)基因多态性芯片检测中的应用。方法　用掺入标记或引物标记荧光分子Cy5 ,扩增HBVP区、前C C区 ,制备含荧光分子Cy5的目的片段后 ,分别与HBV基因多态性芯片杂交、扫描并分析结果。结果　掺入标记法信号强度略高于引物标记法 ,但非特异信号强度也高 ,两法检测 42份乙型肝炎患者血清 ,结果完全一致 ,重复性均较好 ,批内CV 1 5 %～ 2 0 % ,引物标记法用于检测HBV基因多态性灵敏度达 1 0 4copies ml。结论　引物标记法信号强度虽略低于掺入标记法 ,但检测灵敏度。Objective To evaluate the influence of assays with primer labeled with fluorochrome (Cy5) and dUTP labeled with Cy5 on the signal intensity of the chip for detection of hepatitis B virus (HBV) gene polymorphism.Methods The P region and pre C/C region of HBV gene were amplified by polymerase chain reaction (PCR) with Cy5 labeled primer or Cy5 labeled dUTP.The amplicons of the two assays were hybridized with chips,scanned and analyzed by computer software for the detection of HBV gene polymorphism.Results The signal intensity of assay with Cy5 labeled dUTP was slightly higher than that of assay with Cy5 labeled primer,but non specific signal intensity of the assay with Cy5 labeled dUTP was higher.The result of 42 samples showed that there was no significant difference between the two assays,and that both had a good repeatability and CV value (15% 20%).Conclusion The assay with Cy5 labeled primer may replace the assay with Cy5 labeled dUTP as a routine method to detect HBV gene polymorphism,and it is simpler and cheaper.

关 键 词：荧光染料 聚合酶链反应 DNA 基因 多态现象 

分 类 号：R346[医药卫生—基础医学]