| Taqman探针实时定量PCR检测猪伪狂犬病毒 |
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| 引用本文： | 张雪寒,何孔旺,缪文凯,茅爱华,俞正玉.Taqman探针实时定量PCR检测猪伪狂犬病毒[J].江苏农业学报,2008,24(4). |
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| 作者姓名： | 张雪寒 何孔旺 缪文凯 茅爱华 俞正玉 |
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| 作者单位： | 江苏省农业科学院兽医研究所,江苏,南京,210014 |
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| 基金项目： | 国家重点基础研究发展计划(973计划) |
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| 摘 要： | 选取猪伪狂犬病毒(PRV)基因组中gE基因,设计特异性引物和Taqman探针,利用实时定量PCR来定量检测PRV.利用PCR技术扩增猪伪狂犬病毒gE基因80 bp片段,并克隆到pMD18-T载体上,阳性重组质粒命名为pgE80.pgE80质粒进行10倍系列稀释,作为荧光PCR检测的标准模板,定量拷贝数,并绘制标准曲线.以提取的PRV、猪圆环病毒2型(PCV2)、猪细小病毒(PPV)、猪瘟病毒(CSFV)、猪繁殖与呼吸障碍综合症病毒(PRRSV)和传染性胃肠炎病毒(TGEV)的DNA作为模板,进行特异性检测.该实时定量PCR方法,标准曲线斜率为-0.37,R2=0.992,可以检测到20个拷贝数.PRV能被扩增出S型荧光曲线,而其它病毒均未能扩出.用PRV强毒肌肉注射仔猪,定量PCR比普通PCR更能灵敏而特异地检出呼吸道排毒和血液带毒.建立的实时定量PCR方法可用于临床样品快捷、准确、方便地检测.
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| 关 键 词： | 猪伪狂犬病毒 实时定量PCR Taqman探针 定量检测 |
Detection of Pseudorabies Virus by qReal-time PCR with Taqman Probe |
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| ZHANG Xue-han,HE Kong-wang,MIAO Wen-kai,MAO Ai-hua,YU Zheng-yu.Detection of Pseudorabies Virus by qReal-time PCR with Taqman Probe[J].Jiangsu Journal of Agricultural Sciences,2008,24(4). |
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| Authors: | ZHANG Xue-han HE Kong-wang MIAO Wen-kai MAO Ai-hua YU Zheng-yu |
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| Institution: | ZHANG Xue-han,HE Kong-wang,MIAO Wen-kai,MAO Ai-hua,YU Zheng-yu(Institute of Veterinary Medicine,Jiangsu Academy of Agricultural Sciences,Nanjing 210014,China) |
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| Abstract: | A quantitative real-time PCR assay was established for detection of pseudorabies virus.The primers and probe were designed according to the gE gene sequence of pseudorabies virus.The 80 bp region of the PRV gE gene was cloned into pMD18-T vector,and the constructed plasmid was named pgE80.Serial dilutions of plasmid pgE80 were used to quantify the virus genomic copy number,and served as standard curve.PRV,PCV2,PPV,CSFV,PRRSV and TGEV DNA(cDNA) were detected at the same time with the real time qPCR.Results s... |
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| Keywords: | pseudorabies virus quantitative real time PCR Taqman probe quantitative detection |
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