Application of an Improved Real-time TaqMan RT-PCR in FMDV Detection

SHI Li-li; GU Chao-jiang; ZHANG Qian; ZHANG Wei-Ying; LI Yong; LU Bin; HE cheng; QU San-fu; ZHENG Cong-yi*

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October 2006

SHI Li-li, GU Chao-jiang, ZHANG Qian, ZHANG Wei-Ying, LI Yong, LU Bin, HE cheng, QU San-fu and ZHENG Cong-yi*. Application of an Improved Real-time TaqMan RT-PCR in FMDV Detection[J]. Virologica Sinica, 2006, 21(5): 449-453.

Citation: SHI Li-li, GU Chao-jiang, ZHANG Qian, ZHANG Wei-Ying, LI Yong, LU Bin, HE cheng, QU San-fu, ZHENG Cong-yi*. Application of an Improved Real-time TaqMan RT-PCR in FMDV Detection .VIROLOGICA SINICA, 2006, 21(5) : 449-453.

应用改良的实时TaqMan荧光定量RT－PCR技术检测口蹄疫病毒及其3D基因转录水平

1.
武汉大学生命科学学院，病毒学国家重点实验室，湖北武汉430072

摘要

将改良的实时TaqMan荧光定量RT－PCR技术应用于口蹄疫病毒感染体内和体外的定量检测以及其3D基因转录水平分析。结果表明：对样品中口蹄疫病毒基因组RNA的检测灵敏度可达l0个基因拷贝，可同时检测病毒正负链复制水平且重复性较好，所测口蹄疫病毒3D基因转录水平可高达6.9×104拷贝/μL；与实时SYBR GreenⅠ染料RT-PCR技术比较，改良的实时TagMan荧光定量RT－PCR技术检测灵敏度高6.7倍。以上结果证实，改良的实时TaqMan荧光定量RT－PCR技术在病毒检测和基因表达水平分析方面有更高的灵敏度和特异性。
- 口蹄疫病毒
- , 实时TaqMan RT-PCR技术
- , 定量检测

Application of an Improved Real-time TaqMan RT-PCR in FMDV Detection

1.
State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China

Abstract

In this study, an improved real-time RT-PCR was applied to quantification of Foot-and-mouth disease virus in vivo and in vitro. The results suggested that this method had a high sensitivity with up to less than 10 copies / reaction. Furthermore, this novel method was successfully used to quantify the transcriptional level of FMDV-3D gene in transfected BHK-21 cells and the resultant concentration of RNA level was up to 6.9×10 4 copies/μL. Compared with SYBR GreenⅠRT-PCR, six of eight samples testing the FMDV RNA copy number was up to 6.7-fold higher when determined by TaqMan RT-PCR, indicating that the improved method shows a high sensitivity in virus detection.
- Foot-and-mouth disease virus
- , Real-time TaqMan RT-PCR
- , Quantificational detection

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

Application of an Improved Real-time TaqMan RT-PCR in FMDV Detection

1. State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China

Abstract: In this study, an improved real-time RT-PCR was applied to quantification of Foot-and-mouth disease virus in vivo and in vitro. The results suggested that this method had a high sensitivity with up to less than 10 copies / reaction. Furthermore, this novel method was successfully used to quantify the transcriptional level of FMDV-3D gene in transfected BHK-21 cells and the resultant concentration of RNA level was up to 6.9×10 4 copies/μL. Compared with SYBR GreenⅠRT-PCR, six of eight samples testing the FMDV RNA copy number was up to 6.7-fold higher when determined by TaqMan RT-PCR, indicating that the improved method shows a high sensitivity in virus detection.