摘要
本实验通过设计针对伪狂犬病病毒gE基因的引物和MGB（Minor groove binder oligodeoxynucleotide conjugate,MGB-ODN）TaqMan探针,结合ABI PE7700荧光定量PCR仪器系统,建立了一种能区别伪狂犬病病毒野毒株和gE-疫苗株的快速检测方法gE-MGB-TaqMan PCR.实验表明,该方法可检测出最低43拷贝的gE基因和104倍稀释的伪狂犬病病毒Fa株DNA,与微量血清中和结合MTT比色法相比,灵敏度和特异性一致,检测时间仅为后者的1/10,操作比后者更为简单.特异性和重复性试验表明：gE-MGB-TaqMan PCR特异性和重复性好.该方法以闭管的模式操作,减少了各步骤污染的可能性,整个检测少于3 h.
A fluorogenic quantitative PCR was established for detecting and differentiating PRV wild-type strains from vaccine strains depleted of gE gene. The method used a pair of primers specific for gE gene and an internal dual-labeled fluorogenic probe of PRV gE gene based on the AB[ PE7700 detection system. The assay was able to detect 43 copies of gE gene and 104 dilution of PRV Fa strain DNA, with a sensitivity and specificity comparable to that of micro sera neutralization combined with MTT detection. However the PCR method took only 1/10 of the time reqired by MTT and can be performed in less than 3 hours. Also the simplicity of the assay minimized the risk of contamination of subsequent procedures.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2007年第7期550-554,568,共6页
Chinese Journal of Preventive Veterinary Medicine
基金
国家质检总局课题(SK064-02)
