人类疱疹病毒6型荧光定量PCR检测方法的建立和应用  

Establishment and clinical application of TapMan real-time RT-PCR method for detection of HHV-6

作　　者：陈倩倩[1] 张兵[2] 谢志萍[3] 李金松[3] 高寒春[3] 肖霓光[2] 谢乐云[2] 余阗[2] 曾赛珍[2] 龚萍[2] 段招军[1] 

机构地区：[1]南华大学,湖南衡阳421000 [2]湖南省人民医院 [3]中国疾病预防控制中心病毒病预防控制所

出　　处：《中华实验和临床病毒学杂志》2013年第2期144-146,共3页Chinese Journal of Experimental and Clinical Virology

摘　　要：目的建立人类疱疹病毒6型（HHV-6）荧光定量PCR检测方法，并检测临床样本。方法根据文献合成HHV-6U65．66基因片段的特异性引物和TaqMan探针，构建质粒制备标准品，评估该方法的特异性、灵敏度和重复性；并用该方法检测93份临床诊断为病毒性脑炎的脑脊液标本。结果本实验检测HHV-6的灵敏度为3×10^0拷贝／μl；标准曲线间线性关系（R2）为0．999，扩增效率为97．9％；同一样本重复检测3次，组内Ct值的变异系数最大为0．61％，组间为3．13％；特异性检测中只有HHV-6阳性标本出现扩增曲线。93份临床标本中检出HHV-6阳性2例，检出率为2．15％。结论本实验所建立的荧光定量PCR检测HHV-6的方法特异强、灵敏高、重复性好，具有应用于临床检测的潜在价值。Objective To establish a rapid, sensitive and specific real-time PCR method for detection of Human Herpesvirus-6 （HHV-6）. Methods According to the reference,a pair of primers and a probe were designed located in U65-66 gene and to set up the standards. We established a real-time RT- PCR method for detection of HHV-6, and to verify the specificity,sensitivity,reproducibility. Results The correlation coefficient was 0.999, E = 97.9% , the coefficient of variation values of Ct were 0.61% and 3. 13% in real-time PCR assay for inter and intra assay,respectively. The results of all viruses were negative except of HHV-6 for the assay. The quantitative detection limit of the assay was 3 ×10^0copies/μl. Conclusion The real-time PCR assay is highly specific, sensitive and reproducible,which can be used to quatitative detecting clinical samples.

关 键 词：疱疹病毒6型 人 聚合酶链反应 

分 类 号：R4[医药卫生—临床医学] R51