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犬瘟热病毒RT-PCR检测方法的建立
引用本文:郭玲,万莉,马磊,张琦,谢志勇,杨映,舒蕾,陈世界,杨晓农,颜其贵.犬瘟热病毒RT-PCR检测方法的建立[J].中国兽医科学,2012(2):166-170.
作者姓名:郭玲  万莉  马磊  张琦  谢志勇  杨映  舒蕾  陈世界  杨晓农  颜其贵
作者单位:四川农业大学动物医学院;四川出入境检验检疫局;西南民族大学;四川农业大学动物疫病与人类健康四川省重点实验室
基金项目:教育部“长江学者和创新团队发展计划”创新团队项目(IRT0848);成都大熊猫繁育研究基金会项目;四川省科技厅科技支撑计划项目(2010SZ0050);科技部公益性行业科研专项(2009424188)
摘    要:根据GenBank中登录的犬瘟热病毒(CDV)核衣壳蛋白基因序列,设计合成了1对特异性引物,以CDV疫苗株ONP为模板,通过对反应条件进行优化,建立了一种快速检测CDV的RT-PCR方法。结果显示,以该引物进行RT-PCR扩增,能得到与理论设计值大小一致的242bp的特异性条带;该方法最低可以检测出约20pg含量的CDV;对已知CDV阳性和阴性病料进行重复性和稳定性试验,结果均一致;用该方法检测犬细小病毒(CPV)、犬冠状病毒(CCV)、狂犬病病毒(RV)和同为副黏病毒科的新城疫病毒(NDV),均无交叉反应;对32份临床病料进行检测,并与免疫胶体金抗原检测试纸检测结果进行比较,符合率为93.75%。结果表明,此方法特异性强,敏感性高,重复性和稳定性均好,可用于犬瘟热病毒的临床诊断和试验研究。

关 键 词:犬瘟热病毒  RT-PCR  检测

Establishment of RT-PCR method for the detection of canine distemper virus
GUO Ling,WAN Li,MA Lei,ZHANG Qi,XIE Zhi-yong,YANG Ying,SHU Lei,CHEN Shi-jie,YANG Xiao-nong,YAN Qi-gui.Establishment of RT-PCR method for the detection of canine distemper virus[J].Veterinary Science in China,2012(2):166-170.
Authors:GUO Ling  WAN Li  MA Lei  ZHANG Qi  XIE Zhi-yong  YANG Ying  SHU Lei  CHEN Shi-jie  YANG Xiao-nong  YAN Qi-gui
Institution:1,2 (1.College of Veterinary Medicine,Sichuan Agricultural University,Ya’an 625014,China;2.Sichuan Agricultural University/Key Laboratory of Animal Disease and Human Health of Sichuan Province,Ya’an 625014,China; 3.Sichuan Enter-Exit Inspection and Quarantine Bureau,Chengdu 610041,China; 4.Southwest University for Nationalities,Chengdu 610041,China)
Abstract:Based on the published sequence of the nucleocapsid gene of Canine distemper virus(CDV) in GenBank,a pair of specific primers was designed for the detection of CDV.As a template with the CDV-ONP strain,and after optimization of the reaction system,a rapid RT-PCR method was established. In result,a fragment of 242bp,as was expected,was amplified from the reverse transcribed template. As low as 20 pg RNA of CDV could be detected accurately and rapidly. This method had good reproducibility and stability and no cross reaction with canine parvovirus,canine coronavirus,rabies virus and Newcastle disease virus. Suspected disease materials had been tested by RT-PCR,and the RT-PCR results were compared with the immune colloidal gold technique with the coincidence rate up to 93.75%. The results showed that this method may be used for clinical diagnosis and laboratory research.
Keywords:canine distemper virus  RT-PCR  detection
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