| 猪传染性胃肠炎病毒TaqMan荧光定量PCR方法的建立及在疫苗评价中的应用(英文) |
| |
| 引用本文： | 俞正玉,徐向伟,孙冰,何孔旺,郭容利,杜露平,温立斌,张雪寒,茅爱华,倪艳秀,李彬.猪传染性胃肠炎病毒TaqMan荧光定量PCR方法的建立及在疫苗评价中的应用(英文)[J].农业科学与技术,2014(9):1487-1490. |
| |
| 作者姓名： | 俞正玉 徐向伟 孙冰 何孔旺 郭容利 杜露平 温立斌 张雪寒 茅爱华 倪艳秀 李彬 |
| |
| 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,江苏南京210014 |
| |
| 基金项目： | 江苏省农业科技自主创新资金[CX（13）3069]. |
| |
| 摘 要： | 目的]建立检测猪传染性胃肠炎病毒(transmissible gastroenteritis virus,TGEV)的TaqMan荧光定量PCR方法。方法]根据TGEV基因组中保守的N基因序列设计引物和探针,以梯度稀释的含有N基因的重组质粒作为标准品,进行定量PCR反应。结果]该试验建立的荧光定量PCR方法在102~1010拷贝/μl之间具有良好的线性关系,相关系数达到0.99以上,扩增效率在90%~110%之间。该方法的检测灵敏度为10拷贝,敏感性较高,而且特异性较好,与猪的其他病毒核酸均无交叉反应;批内和批间的变异系数低于3%,表明该方法的重复性较好。应用该方法可以对疫苗的免疫效力进行定量检测和评价,也可以对临床病料进行检测。结论]该研究建立的荧光定量PCR方法灵敏度高、特异性好,可为TGEV的流行病学调查、疫苗开发和发病机制等研究提供可靠的工具。
|
| 关 键 词： | 猪传染性胃肠炎病毒 TaqMan荧光定量PCR 检测 |
Development of TaqMan-based Real-time PCR Assay for Detecting Transmissible Gastroenteritis Virus and Its Application in Vaccine Evaluation |
| |
| Zhengyu YU,Xiangwei XU,Bing SUN,Kongwang HE,Rongli GUO,Luping DU,Libin.Development of TaqMan-based Real-time PCR Assay for Detecting Transmissible Gastroenteritis Virus and Its Application in Vaccine Evaluation[J].Agricultural Science ＆ Technology,2014(9):1487-1490. |
| |
| Authors: | Zhengyu YU Xiangwei XU Bing SUN Kongwang HE Rongli GUO Luping DU Libin |
| |
| Institution: | WEN, Xuehan ZHANG, Aihua MAO, Yanxiu NI, Bin LI( Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences / Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing 210014, China) |
| |
| Abstract: | Objective] This study aimed to establish a TaqMan-based real-time PCR assay for detecting transmissible gastroenteritis virus （TGEV）. Method] Primers and a probe were designed according to the conserved sequence of N gene in TGEV genome. After gradient dilution, the recombinant plasmid harboring the N gene was used as a standard for real-time PCR assay to establish the standard curve. Re- sult] The results showed that the established real-time PCR assay exhibited a good linear relationship within the range of 102-10^10 copies/ul; the correlation coefficient was above 0.99 and the amplification efficiency ranged from 90% to 110%. The de- tection limit of real-time PCR assay for TGEV was 10 copies/μl, suggesting a high sensitivity; there was no cross reaction with other porcine viruses, indicating a good specificity; coefficients of variation within and among batches were lower than 3%, suggesting a good repeatability. The established real-time PCR method could be ap- plied in quantitative analysis and evaluation of the immune efficacy of TGEV vac- cines and detection of TGEV in clinical samples. Conclusion] The TaqMan-based real-time PCR assay established in this study is highly sensitive and specific, which can provide technical means for the epidemiological survey of TGEV, development of TGEV vaccines and investigation of the pathogenesis of TGE. |
| |
| Keywords: | Transmissible gastroenteritis virus （TGEV） TaqMan-based real-time PCR： Detection |
| 本文献已被 CNKI 维普 等数据库收录！ |
|
