摘要
根据基因ⅤⅡ型新城疫病毒F基因裂解位点特征设计一对引物和TaqMan探针,建立一种敏感、特异、重复性良好的基因ⅤⅡ型毒株荧光定量RT-PCR鉴别检测方法。以体外转录法制备的cRNA标准品为标准阳性模板,构建标准曲线,并进行各项指标的检测。结果表明,建立的方法特异性良好,最低可检测到102拷贝·μL-1的模板RNA,敏感性比普通RT-PCR高100倍。本试验建立了基因ⅤⅡ型新城疫病毒荧光定量RT-PCR检测方法,且该方法能够鉴别检测常规方法不能区分的新城疫病毒基因ⅤⅡ型强毒株,表明本研究建立的方法可用于此种基因型毒株的快速鉴别检测。
In order to establish a differential diagnosis Real-time RT-PCR with highly sensitivity,specificity and repeatability,a pair of primers and TaqMan probe were designed according to characteristics of cleavage site sequences of F gene of genotype Ⅶ Newcastle disease virus strains in this study.The standard curve was established by using cRNA standard prepared in vitro transcription as positive template,the related indexes were tested.The results showed that this assay has a good effect of differential detection of genotype Ⅶ NDV;102 copies·μL-1 of template RNA could be detected at least.The sensitive of this assay was 100 times higher than conventional RT-PCR.The real-time fluorescent quantitative RT-PCR for detection of genotype Ⅶ Newcastle disease virus was established.This assay could be used as differential diagnosis method of genotype Ⅶ NDV strains,which couldn’t be identified by conventional methods.Our results indicated that the method established in this study can be used as rapid identification assay for the detection of genotype Ⅶ NDVs.
出处
《畜牧兽医学报》
CAS
CSCD
北大核心
2010年第11期1447-1452,共6页
ACTA VETERINARIA ET ZOOTECHNICA SINICA
基金
青岛市科技发展计划项目(07-2-3-5-jch)
关键词
新城疫病毒
基因Ⅶ型
荧光定量RT-PCR
Newcastle disease virus
genotype Ⅶ
real-time fluorescent quantitative RT-PCR
