为了建立猪传染性胃肠炎病毒SYBR-Green Ⅰ荧光定量PCR检测方法,利用RT-PCR技术扩增出猪传染性胃肠炎病毒N基因中324 bp的片段,并克隆到pGEM-T Easy栽体上,纯化的质粒作为模板进行SYBR Green Ⅰ荧光定量PCR扩增并制作标准曲线,建立了猪传染性胃肠炎病毒的荧光定量PCR检测方法.该方法检测灵敏度可达5×101拷贝数·μL-1,与猪伪狂犬病毒、猪蓝耳病毒、猪圆环病毒、猪瘟病毒、猪细小病毒、猪乙型脑炎病毒均不发生交叉反应,具有良好的特异性和重复性.结果表明,建立的实时荧光定量PCR具有特异、敏感、快速、定量、重复性好等优点,可用于临床TGEV N基因的检测.%To develop the SYBR Green I real-time quantitative PCR assay for detection of N gene of transmissible gastroenteritis virus ( TGE V ) . A 324 bp region of the transmissible gastroenteritis virus N gene was amplified using reverse transcriptase polymerase chain reaction (RT-PCR) assay, then subcloned to pGEM-T vector and acquired the recombinant plasmid pGEM-N, which served as template to conduct the standards curve of the SYBR Green I real-time PCR. The results showed that the SYBR Green I real-time PCR was established and could used to detect TGEV. The TGEV sensitivity analysis showed that the developed SYBR Green I real-time PCR could detect 5 x 10' copy ·μL-1. And the specificity and repeatability of this method were very good. All the results suggested that the SYBR Green I real-time PCR we established in present study could be used in clinical diagnosis and epidemiological investigation for TGEV.
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机译：Development of TaqMan-based Real-time PCR Assay for Detecting Transmissible Gastroenteritis Virus and lts Application in Vaccine Evaluation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
