中国媒介生物学及控制杂志

• 论著 • 上一篇    下一篇

应用荧光定量PCR检测环境样品中鼠疫耶尔森菌

李伟;海荣;俞东征;张志凯;蔡虹;张建华   

  1. 中国疾病预防控制中心传染病预防控制所人兽共患病研究室 北京
  • 出版日期:2005-08-20 发布日期:2005-08-20

Detection of Yersinia pestis from Mice Tissues and Soil by Realtime Fluorescence Polymerase Chain Reaction

LI Wei;HAI Rong;YU Dong-zheng; et al   

  1. Institute for Communicable Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing 102206,China
  • Online:2005-08-20 Published:2005-08-20

摘要: 目的应用荧光定量聚合酶链反应(PCR)法检测土壤和动物脏器中鼠疫耶尔森菌,评价并优选环境样品目标核酸的纯化方法。方法应用TaqMan荧光定量PCR技术,以鼠疫耶尔森菌placaf1hms引物、探针为指标,比较不同种从土壤和脏器中纯化鼠疫耶尔森菌核酸的方法。结果应用NaI-Glassmilk方法及荧光定量PCR检测土壤中的鼠疫菌,检出底限为104拷贝/g土壤(pla),从染疫动物脾脏、肝脏标本中也能敏感检出鼠疫菌。结论荧光定量PCR方法和NaI裂解Glassmilk制备模板联合应用可灵敏特异地从土壤和动物脏器中检测鼠疫耶尔森菌。

关键词: 鼠疫耶尔森菌, 土壤, 动物脏器, 荧光定量PCR

Abstract: Objective To establish and assess different methods for detection of Yersinia pestis from artificially contaminated soil and infected mice tissues by realtime fluorescence polymerase chain reaction. Methods Different methods for isolation and purification of nucleic acids from soil and mice tissues contaminated by Y.pestis vaccine strain were established. Three combinations of primers and probes designed according to caf1 and pla and hms genes of Y.pestis were used to assessed these methods. The optimal systems were confirmed by comparing the amplification results. Results The method of NaIGlassmilk is the most effective,and ten thousand of bacteria in one gram soil could be directly detected by this system. Conclusion The NaIGlassmilk method and the Realtime fluorescence polymerase chain reaction could directly and rapidly detect Y.pestis in contaminated issues or soil.