【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10-4,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。更多还原

【Abstract】 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 for 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.更多还原