动物源性成分实时等温扩增检测方法的建立  

Loop-mediated isothermal amplification assay for identification of animal derived materials

作　　者：宋涛平 杨丽霞 邱华丽 邱志鹏 SONG Tao-Ping YANG LI-Xia QIU Hua-Li QIU Zhi-Peng(Changsha Institute for Food and Drug Control, Changsha 410016, China Food Safety Production Engineering Research Center of Hunan Province, Changsha 410016, China)

机构地区：[1]长沙市食品药品检验所,长沙410016 [2]湖南省食品安全生产工程技术研究中心,长沙410016

出　　处：《食品安全质量检测学报》2017年第8期3207-3212,共6页Journal of Food Safety and Quality

基　　金：长沙市科技计划项目(kh1601093)~~

摘　　要：目的建立一种实时荧光环介导等温扩增法(loop-mediated isothermal amplification,LAMP)快速检测动物源性成分的分析方法。方法根据鸡、鸭、牛、羊物种cytb、COX3、12S rRNA、cytb特异性基因分别设计LAMP引物,反应时加入荧光染料SYTO-9,在63℃条件下反应45min,通过实时检测荧光强度判断反应结果,并进行实时监测。结果实时荧光LAMP方法对鸡、鸭源性成分的检测灵敏度为1pg,对牛、羊源性成分的检测灵敏度为10pg。对熟肉制品中掺杂肉检测时,掺杂鸡肉、鸭肉、牛肉和羊肉的检出限为0.01%。结论该方法可用于快速检测肉制品中动物源性成分,具有操作简单、无需贵重仪器、反应过程可实时监控、灵敏度高和特异性强等优点,适合于基层监管部门的现场快速检测。Objective To establish a rapid real-time loop-mediated isothermal amplification （LAMP） method for rapid detection of animal derived materials. Methods The specific LAMP primers of chicken, duck, cattle and sheep were designed according to the specific mitochondrial cytb, COX3, 12S rRNA and cytb genes, respectively. Target DNA was amplified by real-time LAMP with fluorescent dye SYTO-9 within 45 min at an isothermal temperature of 63 ℃. The reaction results were judged by detecting the fluorescence intensity in real-time, and real-time monitoring was carried out. Results The detection limits of LAMP assay were 1 pg genomic DNA for chicken and duck and 10 pg for cattle and sheep.Thedetectionlimitsforadulteratedmeatproductscouldreach0.01%forchicken,duck,cattleandsheep. Conclusion This method for rapid detection of meat products has the advantages of simplicity, low cost, real-time monitoring, high sensitivity and specificity, and is suitable for rapid on-site detection of supervision departments.

关 键 词：动物源性成分 环介导等温扩增 检测 

分 类 号：TS251.7[轻工技术与工程—农产品加工及贮藏工程]