牛新孢子虫内标多重PCR检测方法的试验  

Study on multiplex PCR with internal control for detection of Neospora caninum

作　　者：唐慧芬[1,2] 季新成[2] 于学辉[2] 

机构地区：[1]石河子大学生命科学学院,新疆石河子832000 [2]新疆出入境检验检疫局,新疆乌鲁木齐830063

出　　处：《中国兽医杂志》2012年第4期23-26,共4页Chinese Journal of Veterinary Medicine

基　　金：国家质量监督检验检疫总局科研项目(2009IK011;2011IK017)

摘　　要：本试验以牛新孢子虫NcSAG1基因、Nc-5基因和NcSRS2基因作为检测新孢子虫病的目的基因,以牛性腺(prolac-tin)基因作为内标对反应过程进行监测,对上述基因各设计1对引物。结果表明,4对引物可分别扩增出201bp、231bp、256bp和156bp的目的条带。经反应条件的优化,该方法具有较好的灵敏度,各质粒在103 copies/反应时,均可于同一管中扩增出较清晰的目的条带,只比单重PCR的灵敏度低了10倍,且不受内标模板存在的影响。经临床应用研究,该方法具有较好的特异性和重复性,可用来对新孢子虫进行快速准确的检测。Using NcSAG1,Nc-5 and NcSRS2 gene as target genes for the detection of Neospora caninum and prolactin gene as internal control to monitor reaction process,four pairs of primers were designed.The results showed that the four pairs of primers amplified four fragments of 201 bp,231 bp,256 bp and 156 bp in length,respectively.The detection limit was 103 copies/reaction after optimization of reaction conditions,which was only 10 times lower than single PCR.The sensitivity of multiplex PCR was not affected by the existence of the intenal control.The method had higher specificity and reliability,and could be used to detect Neospora caninum rapidly and accurately.

关 键 词：新孢子虫病 多重PCR 内标 共扩增 

分 类 号：S852.723[农业科学—基础兽医学]