一种产耐热蛋白酶假单胞菌双重PCR检测方法的建立  

Development of a Duplex PCR Method for Detecting Proteolytic Pseudomonas

作　　者：牛春华[1] 赵玉娟[1] 高磊[1] 段翠翠[1] 李盛钰[1] 

机构地区：[1]吉林省农业科学院农产品加工研究所国家乳品加工技术研发分中心,吉林长春130033

出　　处：《农产品加工（下）》2015年第9期43-46,共4页Farm Products Processing

基　　金：吉林省世行贷款农产品质量安全项目(2011-Y35);现代农业产业技术体系专项资金资助(CARS-37)

摘　　要：假单胞菌是影响原料乳品质的主要嗜冷菌,拟建立一种针对假单胞菌及其耐热蛋白酶的双重PCR检测方法。针对假单胞菌的16S r DNA及apr X基因设计多对引物,利用单PCR反应进行筛选。利用优选的2对引物进行双重PCR反应体系优化,PCR产物的测序鉴定和特异性试验,建立了一种能同时检测分泌蛋白酶的假单胞菌双重PCR方法。用此方法对假单胞菌、肠道常见致病菌进行双重PCR检测,结果表明仅具有蛋白水解活性的假单胞菌在16S r DNA和apr X基因扩增区得到有效扩增,而致病菌和阴性对照在两扩增区均为阴性,表明此检测方法具有特异性。Pseudomonas is a kind of key psychrophilic bacteria affecting the quality of raw milk. This study aimed to establish a duplex PCR method to detect the pseudomonas and thermophilic protease. Several pairs of specific primers are designed for 16S rRNA gene and aprX gene of Pseudomonas according to the gene sequences from GenBank, and PCR method is used to screen proper primers. Two pairs of selected primers are used to optimize the reaction conditions of duplex PCR. Meanwhile, this method is used to detect the Pseudomonas and common intestinal pathogenic bacterium. The results show that only the 16S rDNA and aprX genes from Pseudomonas with proteolytic activity could be amplified. It can be concluded that the duplex PCR is specific and could be used in detecting proteolytic Pseudomonas.

关 键 词：假单胞菌 蛋白水解 双重PCR 

分 类 号：TS252.7[轻工技术与工程—农产品加工及贮藏工程]