应用TaqMan荧光定量RT-PCR检测猪肝中戊型肝炎病毒的研究

doi:10.13217/j.scjpm.2015.0171

华南预防医学 ›› 2015, Vol. 41 ›› Issue (2): 171-175.doi: 10.13217/j.scjpm.2015.0171

应用TaqMan荧光定量RT-PCR检测猪肝中戊型肝炎病毒的研究

林虹^1，2，管大伟²，邓小玲^1，3，4，梁辉^3，4，李柏生²，周少君^3，4，钟贤武^1，2，柯昌文²

1.中山大学公共卫生学院，广东广州 510080；2.广东省疾病预防控制中心广东省应急病原学重点实验室；3.广东省疾病预防控制中心；4.国家食品安全风险监测广东中心

出版日期:2015-04-25 发布日期:2015-05-06
通讯作者: 邓小玲 E-mail:2296895035@qq.com
作者简介:林虹（1988—），女，硕士在读，研究方向：食源性疾病分子流行病学
基金资助:
广东省病原微生物种资源保藏与利用（No. 2012B06040012）；十二五重大科技专项—病原谱研究（No. 2012ZX10004213-004）

Application of TaqMan based real-time fluorescent quantitative RT-PCR assay in detection of hepatitis E virus in swine liver

LIN Hong^1,2, GUAN Da-wei², DENG Xiao-ling^1,3,4, LIANG Hui^3,4, LI Bo-sheng², ZHOU Shao-jun^3,4, ZHONG Xian-wu^1,2, KE Chang-wen²

1.School of Public Health, Sun Yat-sen University, Guangzhou 510080, China; 2.Guangdong Provincial Center for Disease Control and prevention, Guangdong Provicial.Key Laboratory of Pathogen Detection fer Emergency Response; 3.Guangdong Provincial Center for Disease Control and prevention; 4.Guangdong Center for National Food Safety Risk Surveillance

Online:2015-04-25 Published:2015-05-06

摘要/Abstract

摘要： 目的建立适用于猪肝样本中戊肝病毒检测的荧光RT-PCR方法。方法采用匀浆处理直接提取法（方法一）与聚乙二醇处理法（方法二）两种处理方法对染毒猪肝样本进行病毒富集，针对4种型别戊肝病毒的基因组而设计的特异性引物和探针，建立一套TaqMan荧光定量RT-PCR方法，并评价该方法的准确性和灵敏性。采用新建荧光定量RT-PCR检测人工染毒的猪肝样本并比较两种方法的处理效率。结果所建立的TaqMan荧光定量RT-PCR标准曲线显示在102～108 copies/μL范围内具有良好的线性关系，相关系数为0.999。批内和批间重复性实验结果表明，不同浓度的质粒标准品检测所获得的Ct值的变异系数分别在0.11%～3.75%、2.47%～6.62%间。2种方法均可以检测出102 copies/μL的染毒猪肝中戊肝病毒拷贝数。结论所建立的TaqMan荧光定量RT-PCR方法具有灵敏度高、稳定性好、特异性强等特点，适用于HEV的定量检测。匀浆处理直接提取法更加快捷，适用于长期监测。

Abstract: Objective To develop a simple, quick, and accurate detection method for distinguishing hogwash oil from raw vegetable. Methods Testing samples included 106 raw vegetable oils in 10 categories and 11 batches of refined hogwash oils. The Raman spectra were scanned to compare the morphology and acquire the first derivatives, including the lower limit value for each kind of raw vegetable oils to establish steps of the method. The accuracy was evaluated by detecting 234 mixture samples. Results There was morphology of smooth bulk peak in all refined hogwash oils, and the 2 839 cm-1 peak values were negative for the first derivatives. The ratios of peak 2 944 to 2 839 cm-1 were between -0.851 and -3.015. The accuracies for the mixtures with 10% and 20% hogwash oil were 80% and 100%, respectively. ConclusionRaman spectra of hogwash oil and raw plant oil were significantly different and can be used as reference for distinguishing between them.

中图分类号:

R440

林虹，管大伟，邓小玲，梁辉，李柏生，周少君，钟贤武，柯昌文. 应用TaqMan荧光定量RT-PCR检测猪肝中戊型肝炎病毒的研究[J]. 华南预防医学, 2015, 41(2): 171-175.

LIN Hong, GUAN Da-wei, DENG Xiao-ling, LIANG Hui, LI Bo-sheng, ZHOU Shao-jun, ZHONG Xian-wu, KE Chang-wen. Application of TaqMan based real-time fluorescent quantitative RT-PCR assay in detection of hepatitis E virus in swine liver[J]. S China J Prev Med, 2015, 41(2): 171-175.

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应用TaqMan荧光定量RT-PCR检测猪肝中戊型肝炎病毒的研究

Application of TaqMan based real-time fluorescent quantitative RT-PCR assay in detection of hepatitis E virus in swine liver

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