框镜鲤致病性维氏气单胞菌双重PCR检测方法的建立  

Development of duplex PCR for detection of Aeromonas veronii

作　　者：边宇[1] 钱宏伟 孟庆峰[3] 贺德聪 比尔来西肯.赛都力 单晓枫[1] 康元环[1] 王伟利[3] 钱爱东[1] 

机构地区：[1]吉林农业大学动物科学技术学院,吉林长春130118 [2]吉林电力医院,吉林长春130022 [3]吉林出入境检验检疫局,吉林长春130062 [4]吉林省通化市畜牧总站,吉林通化134000 [5]伊犁职业技术学院,新疆伊犁835000

出　　处：《中国预防兽医学报》2013年第4期304-307,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家支撑计划项目(2010BAD04B01);吉林省科技厅项目(20080218);吉林农业大学青年启动基金(200904)

摘　　要：为建立快速检测框镜鲤致病性维氏气单胞菌(A.veronii)双重PCR方法,本研究以A.veronii CY0806株和国际标准株DNA为模板,分别以16S rRNA和Aer基因特异性引物进行PCR扩增,分别获得大小约880 bp和430 bp的DNA片段。通过序列比对分析,16S rRNA基因片段、Aer片段序列与GenBank中登录的A.veronii ATCC35624株的的同源性均为99%。进一步试验显示该方法的敏感性较高,达到1.58×10-3ng/μL,特异性较强,只有A.veronii标准株及分离株结果呈阳性;人工模拟污染样本试验显示:该方法的检出率达到了86.7%,高于细菌分离培养的70%检出率。双重PCR检测方法的建立,为框镜鲤致病性A.veronii的检测提供新的方法。To establish the duplex PCR method for detection of Aeromonas Veronff in Cyprinus carpio, two DNA fragments about 880 bp and 430 bp were amplified with the specific primers of 16S rRNA gene and Aergene from the chromosome DNA of A. veronii CY0806 strain and reference strain. Alignment analysis indicated that the 2 DNA fragments shared 99% homology in sequence with the gene of A.veronii ATCC35624. The further experiments indicated that the duplex PCR method was specific for A.veronii with a limit detection of 1.58x103 ng bacteria DNA. In addition, tested on 30 artificial infected fish samples showed that the detection rate of the duplex PCR method reached 86.7% which was higher than that of 70% by bacterial isolation method. The establishied duplex PCR provided a sensitive method for the detection of A. veronii infection in Cypnnus carpio.

关 键 词：维氏气单胞菌 气溶素基因 16S RRNA 双重PCR 

分 类 号：S852.61[农业科学—基础兽医学]