不同引物检测甲型H1N1流感病毒的比较研究  

Study on the sensitivity of different primers at detecting nucleic acids of the influenza A H1N1 virus

作　　者：李忠[1] 韩莹 刘倜[1] 张圣洋[1] 王宇路[1] 王显军[1] 徐爱强[1] 毕振强[1] 

机构地区：[1]山东省疾病预防控制中心,山东省传染病预防控制重点实验室,山东大学预防医学研究院,山东济南250014 [2]山东省艾克韦生物技术有限公司

出　　处：《中国病原生物学杂志》2011年第4期262-264,共3页Journal of Pathogen Biology

基　　金：山东省科技厅计划项目(No.2009GG10002054)

摘　　要：目的筛选灵敏、特异的甲型H1N1流感病毒核酸检测方法。方法采用国家流感中心推荐的甲型流感病毒（H1N1）核酸引物和1对自行设计的引物，同时选择市售荧光定量PCR试剂盒，分别对不同浓度的流感病毒进行RT-PCR或荧光定量PCR扩增，比较其检测的灵敏度和特异性。结果荧光定量PCR检测H1N1病毒核酸的灵敏度为10^-5（病毒稀释度），高于普通RT-PCR的灵敏度10^-3～10^-4；RT-PCR扩增甲型流感病毒（H1N1）核酸时，自行设计引物的灵敏度为10，高于中国CDC推荐引物的灵敏度10^-3。2种引物的特异性一致。结论对于甲型（H1N1）流感病毒疑似样品的检测，荧光定量PCR是较灵敏的方法，但从成本和灵敏度两方面考虑，自行设计的引物更适合基层疾控机构采用。Objective To select an assay with high sensitivity and specificity at detecting nucleic acids of the influenza A H1N1 virus. Methods Sensitivity was analyzed using RNA extracted from a 10-fold serially diluted viral culture. Nucleic acids were amplified using fluorescence quantitative PCR and RT-PCR. Two pairs of primer were used for RT-PCR. One pair of primers was designed by the authors and the other was recommended by the Chinese Center {or Disease Control and Prevention. Results Fluorescence quantitative PCR was more sensitive than RT-PCR. The detection limit of fluorescence quantitative PCR was found to be 10^-5 dilution, while the detection limit of RT-PCR was 10^-3 or 10^-4 dilution. The detection limit of RT-PCR using the primers that the authors designed was 10^-4, while that of RT-PCR using the primers designed by the Chinese Center for Disease Control and Prevention was 10^-3. There was no difference in the specificity of these two pairs of primers for RT-PCR. Conclusion Fluorescence quantitative PCR was found to be the most sensitive method of detecting suspected cases of influenza A H1N1. In light of their cost and sensitivity, the primers synthesized by the authors are the most suitable.

关 键 词：甲型流感(H1N1) 引物 灵敏度 

分 类 号：R383.24[医药卫生—医学寄生虫学]