摘要
根据GenBank中的布鲁菌属特异性基因序列BCSP31设计1对引物,以牛种布鲁菌544A、牛种布鲁菌104M、羊种布鲁菌16M和猪种布鲁菌S2基因组DNA作为模板,进行PCR扩增。优化PCR的反应条件,并对方法的特异性、敏感性及适用性进行验证。结果显示,PCR可扩增得到287 bp的目的基因条带,测序结果与已发表的序列同源性达100%。该方法对牛种布鲁菌544A的最小检出量为0.16 pg,对大肠杆菌O157∶H7、小肠结肠炎耶尔森菌等15种参照菌的核酸扩增结果均为阴性。建立的PCR方法具有快速简便、特异性强、敏感性高等特点,为布鲁菌的实验室诊断和流行病学调查奠定了基础。
A pair of primers were designed according to the sequence of specific Brucella genus gene BCSP31 reported in GenBank,amplified using the bacterial genome of B.abortus544A,B.abortu104M,B.melitensis16M and B.suisS2 strains as a template,the reaction condition for PCR was optimized,and specificity,sensitivity and adaptivity of the developed method were verified.The target gene band at a length 287 bp was amplified by PCR,with a homology of 100% to the gene sequence reported in GenBank.The PCR assay could detect as low as 0.16 pg of Brucella abortus 544A DNA.All the detection results of fifteen non-Brucella species by the developed method was negative.A rapid,specific and sensitive PCR method for detection of Brucella was developed.This protocol can be used for brucellosis diagnosis in laboratory and contributed to epidemiological investigation of brucellosis.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2010年第11期1486-1489,1499,共5页
Chinese Journal of Veterinary Science
基金
吉林省省长基金资助项目(20070613)
国家科技支撑计划资助项目(2008BAD96B11)
