摘要
为建立犬瘟热病毒(CDV)SYBR Green Ⅰ实时荧光定量PCR检测方法,采用RTPCR扩增CDV F基因269bp片段,并克隆入pMD18-T载体中,以纯化的重组质粒为模板作荧光定量PCR扩增,建立了CDV荧光定量PCR检测方法。该方法检测灵敏度可达1.6×101拷贝/μL,与犬细小病毒(CPV)、犬腺病毒(CAV-1)、犬冠状病毒(CCV)、犬副流感病毒(CPIV)均不发生交叉反应,具有很好的特异性和重复性。结果表明:建立的CDV实时荧光定量PCR具有特异、敏感、快速、定量、重复性好等优点,可用于临床犬瘟热(CD)的检测。
To develop the SYBR Green I real-time quantitative PCR assay for detection of F gene of Ca- nine Distemper Virus（CDV）, a 269bp conservative region from CDV F gene was amplified by RT-PCR and cloned into pMD18-T vector to construct recombinant plasmid of pMD18-VP60, which served as a template to establish the standard curve of the SYBR Green I realtime PCR. The results showed that the SYBR Green I real-time PCR was specifically detected by CDV with a limited detection of 1.6 ×101 copy/μL and no amplification for Canine parvovirus, hepatitis infectiosa canis virus, Canine coronavirus and Canine was detected and a good specificity and repeatability was observed. There data suggested that the SYBR Green I real-time PCR which has the merits of specificity, sensitivity, rapidity, fixed quantity and good repeat- ability could be applied to clinical diagnosis and epidemiological investigation for CDV.
出处
《家畜生态学报》
北大核心
2014年第3期58-62,共5页
Journal of Domestic Animal Ecology
