改良LAMP检测鸡肉中单增李斯特氏菌的研究  

Study on Improved Loop-mediated Isothermal Amplification Method for Detection of L. monocytogenes

作　　者：冯显显 杨倩 张先舟[1] 马晓燕[1] 李英军[1] 张伟[1,2] 王建昌 陈启跃 FENG Xian-xian;YANG Qian;ZHANG Xian-zhou;MA Xiao-yan;LI Ying-jun;ZHANG Wei;WANG Jian-chang;CHEN Qi-yue(College of Food Science and Technology,Hebei Agricultural University,Baoding 071001,Hebei,China;College of Science and Technology,Heibei Agricultural University,Cangzhou 061100,Hebei,China;Hebei Entry-Exit Inspection and Quarantine Bureau Inspection and Quarantine Technical Center,Shijiazhuang 050051,Hebei,China;Beijing Jinnomei Biotechnology Co.,Ltd.,Beijing 100176,China)

机构地区：[1]河北农业大学食品科技学院,河北保定071001 [2]河北农业大学理工学院,河北沧州061100 [3]河北出入境检验检疫局检验检疫技术中心,河北石家庄050051 [4]北京金诺美生物技术有限公司,北京100176

出　　处：《食品研究与开发》2018年第19期130-136,共7页Food Research and Development

基　　金：河北省自然科学基金面上项目(C2008000216);河北省高等学校科学技术研究重点项目(ZD2017237)

摘　　要：本研究建立一种改良环介导等温扩增(loop-mediated isothermal amplification,LAMP)技术检测鸡肉中单增李斯特氏菌(Listeria monocytogenes,L. monocytogenes)的方法。针对单增李斯特氏菌的溶血素基因(hlyA)设计LAMP引物,对单增李斯特氏菌进行特异性检测,通过荧光曲线和肉眼观察荧光颜色来判定检测结果。试验结果表明:改良LAMP方法检测单增李斯特氏菌具有良好的特异性,7株单增李斯特氏菌呈阳性结果,24株非单增李斯特氏菌呈阴性结果。与聚合酶链反应(polymerase chain reaction,PCR)方法相比,改良LAMP方法具有灵敏度高(5.4×100 fg/μL)、检出限低(3.9×100 CFU/g)的特点,均是PCR结果的100倍。对66份鸡肉样品进行检测,得到改良LAMP方法的敏感性为100%,特异性为98.41%,符合率为98.48%。综上所述,改良LAMP方法能够快速、准确的检测单增李斯特氏菌,具有很好的应用前景。In order to detect Listeria monocytogenes(L.monocytogenes),an improved loop-mediated isothermal amplification(improved LAMP)detection method was established.The hemolysin(hlyA)gene of L.monocytogenes was used to design the primers of improved LAMP and the detection results were shown by fluorescence curve or the fluorescence color by naked eye.7 strains of L.monocytogenes were positive results and 24 strains of non-L.monocytogenes were negative results.The results indicated that the improved LAMP had a good specificity.Compared with Polymerase chain reaction(PCR),the improved LAMP method had a high sensitivity(5.4×100 fg/μL)and a low detection limits(3.9×100 CFU/g),which were both 100-folds of the results of PCR.The sensitivity,specificity and the coincidence rate of the improved LAMP method were 100%,98.41%and 98.48%,respectively,by detecting 66 chicken samples.In conclusion,it is rapid and accurate for the detection of L.monocytogenes using improved LAMP method which has a good application prospect.

关 键 词：改良环介导等温扩增方法 单增李斯特氏菌 hlyA基因 检测 鸡肉 

分 类 号：TS251.55[轻工技术与工程—农产品加工及贮藏工程]