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王宇, 任莉娜, 杜溪乔, 张炳丽, 宋晓琳. 水体中GII型诺如病毒荧光定量RT-PCR检测[J]. 中国公共卫生, 2016, 32(10): 1441-1444. DOI: 10.11847/zgggws2016-32-10-38
引用本文: 王宇, 任莉娜, 杜溪乔, 张炳丽, 宋晓琳. 水体中GII型诺如病毒荧光定量RT-PCR检测[J]. 中国公共卫生, 2016, 32(10): 1441-1444. DOI: 10.11847/zgggws2016-32-10-38
WANG Yu, REN Li-na, DU Xi-qiao.et al, . Detection of waterborne norovirus with realtime fluorescense quantitative RT-PCR[J]. Chinese Journal of Public Health, 2016, 32(10): 1441-1444. DOI: 10.11847/zgggws2016-32-10-38
Citation: WANG Yu, REN Li-na, DU Xi-qiao.et al, . Detection of waterborne norovirus with realtime fluorescense quantitative RT-PCR[J]. Chinese Journal of Public Health, 2016, 32(10): 1441-1444. DOI: 10.11847/zgggws2016-32-10-38

水体中GII型诺如病毒荧光定量RT-PCR检测

Detection of waterborne norovirus with realtime fluorescense quantitative RT-PCR

  • 摘要: 目的 建立水体中诺如病毒的富集和荧光定量RT-PCR检测方法。方法 梯度稀释诺如病毒阳性便样标本,模拟水体通过含有阳离子混合醋酸纤维膜的微生物抽滤系统,加入浓度20%的PEG6000,高速离心2次浓缩水体获取病毒粒子,以构建标准质粒作对照,荧光定量RT-PCR检测水体中GII型诺如病毒。结果 重组质粒T-NVS定量荧光PCR扩增良好,线性相关系数98%,最低检测灵敏度为100 copies/μL;10-1~10-4倍稀释的模拟水样经富集浓缩后,病毒回收率为5.75~19.74%。结论 初步建立了水体中诺如病毒的富集与检测方法。

     

    Abstract: Objective To establish a method for waterborn norovirus enrichment and detection with realtime reverse transcription(RT)-PCR.Methods Norovirus positive fecal samples were diluted gradiently with purified water;the simulated water samples were filtered through a microbial purification system with cation mixed cellulose acetate membrane and added with 20%PEG6000;then the virions in the samples were obtained with two times of high-speed centrifugation and the nucleic acids of the virus were extracted and detected with fluorescence quantitative realtime RT-PCR.Results Fluorescence quantitative realtime RT-PCR revealed a good amplification of recombinant plasmid (T-NVS)with a linear correlation of 98%;the detection limit of the established method for norovirus was 100 copies/μL.After the enrichment,the virus recovery rate ranged form 5.75% to 19.74% for the simulated water samples with the dilution ratios of 10 to 1 000.Conclusion A method for enrichment and detection of norovirus in water samples was preliminarily established.

     

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