| 猪瘟病毒Taqman实时定量RT-PCR检测方法的建立和临床应用 |
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| 引用本文： | 毛立,李文良,李彬,江杰元.猪瘟病毒Taqman实时定量RT-PCR检测方法的建立和临床应用[J].江苏农业学报,2012,28(4):783-786. |
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| 作者姓名： | 毛立 李文良 李彬 江杰元 |
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| 作者单位： | 江苏省农业科学院兽医研究所,农业部兽用生物制品工程技术重点实验室,国家兽用生物制品工程技术研究中心,江苏南京210014 |
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| 基金项目： | 江苏省农业科技自主创新基金项目 |
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| 摘 要： | 根据猪瘟病毒5’非编码区(5’-UTR)设计特异性引物和Taqman探针,建立Taqman实时定量RT-PCR检测猪瘟病毒法。检测结果显示,该方法的灵敏度为1μl 10拷贝,在病毒拷贝数为1μl 108～101时,循环数(Ct)值与拷贝数对数呈现较好的线性关系,且重复性好,批间变异系数小于1%。用该方法检测猪繁殖与呼吸综合征病毒、牛病毒性腹泻病毒,结果均为阴性。用该方法检测采集自江苏和新疆的192份组织和血清样品,猪瘟病毒阳性率为71.9%;检测感染猪的不同脏器,发现在心、肺、肝、肾、脑、脾脏、淋巴结、腹水中均可以检测到猪瘟病毒,与常规RT-PCR方法相比,该方法敏感性更高。该方法的建立为猪瘟病毒的流行病学调查和定量提供了有效手段。
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| 关 键 词： | 猪瘟病毒 实时定量RT-PCR 临床应用 |
Development and clinical application of Taqman real-time RT-PCR assay for detection of classical swine fever virus |
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| MAO Li
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LI Wen-liang
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LI Bin
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ANG Jie-yuan.Development and clinical application of Taqman real-time RT-PCR assay for detection of classical swine fever virus[J].Jiangsu Journal of Agricultural Sciences,2012,28(4):783-786. |
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| Authors: | MAO Li LI Wen-liang LI Bin ANG Jie-yuan |
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| Institution: | (Institute of Veterinary Medicine,Jiangsu Academy of Agricultural Sciences/Key Laboratory of Veterinary Biological Engineering and Technology,Ministry of Agriculture/National Center for Engineering Research of Veterinary Bio-products,Nanjing 210014,China) |
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| Abstract: | According to the conservative sequences located on the 5′ untranslated region(5′-UTR) of classical swine fever virus(CSFV),a pair of specific primers and Taqman probe were designed and synthesized respectively,and a Taqman real-time fluorescent quantitative reserve-transcribed polymerase chain reaction(real-time RT-PCR) for detecting the CSFV was established in this study.Test results showed that the method had a detection limit of 10 copies of target RNA per reaction,and there was a good linear relationship between Ct value and copy numbers in diluted samples.The variation between batches was less than 1%.The RNA of porcine reproductive and respiratory syndrome virus,bovine viral diarrhea virus were detected by the Taqman RT-PCR,and the results were all negative.The CSFV-positive rate was 71.9% in 192 samples collected from Jiangsu and Xinjiang areas.Real-time RT-PCR detection showed that the different organs of swine including hearts,lungs,livers,kidneys,brains,spleens,lymph nodes and ascites were CSFV-positive,indicating that the method were more sensitive and effective than traditional RT-PCR. |
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| Keywords: | classical swine fever virus(CSFV) Taqman real-time RT-PCR clinical application |
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