饲料中狐狸、水貂、貉子和狗源性的五重实时荧光PCR检测方法的建立  

Multiplex Fluorescent Real-time PCR Detection of Fox,Mink,Raccoon and Dog Derived Materials in Feedstuff

作　　者：刘艳艳[1] 李会荣 胡悦 范阳阳[1] 李祥明 谭晴晴[1] 吴家强[3] 步迅[1] 

机构地区：[1]山东省农业科学院生物技术研究中心,济南250100 [2]山东省饲畜牧兽医局饲料质量检验所,济南250022 [3]山东省农业科学院家禽所,济南250100

出　　处：《中国生物工程杂志》2017年第12期67-76,共10页China Biotechnology

基　　金：国家自然基金青年科学基金(31702215);泰山学者项目;山东省现代农业产业技术体系(SDAIT)资助项目

摘　　要：为了快速鉴别饲料中的狐狸、水貂、貉子和狗源性成分,根据线粒体16S r DNA种间保守序列,设计合成针对狐狸、水貂、貉子和狗的特异性引物和探针,通过对荧光PCR反应体系和反应条件的优化筛选,建立了多重实时荧光PCR方法,在同一PCR反应体系中可以同时完成4种动物源性成分检测。通过对15种其他物种的源性成分的检测,结果表明所设计的引物和探针具有很好的物种特异性,且灵敏度高,狐狸、水貂、貉子和狗的DNA检出限为0.01ng。对40份样品检测,其中5份检测出貉子、狐狸和水貂源性成分。结果表明,该方法可以有效地鉴别出饲料中狐狸、水貂、貉子和狗源性成分,同时适用于相关动物产品中。In order to identify the fox, mink, raccoon and dog-derived components in feed, the specific primers and probes for fox, mink, raccoon and dog were designed and synthesized according to the conserved sequence of mitochondrial 16S rDNA. Fluorescent PCR reaction system and reaction conditions were optimized. A number of real-time fluorescent PCR methods were established. Four kinds of animal-derived components were detected in the same PCR reaction system. The developed method could successfully detect four kinds of animal derived materials including fox, mink, raccoon and dog in the same reaction. Through the detection of the source components of 15 other species, the results showed that the primers and probes designed in this experiment had good species specificity. The method has high sensitivity, fox, mink, raccoon and dog DNA detection limit of 0.01ng. Of the 40 samples tested, 5 of them detected raccoon, fox and mink-derived ingredients. The results showed that the method has high specificity and sensitivity. It could effectively identify the fox, mink, raccoon and dog - derived components in the feed, and it was suitable for the related animal products.

关 键 词：线粒体16S r DNA多重实时荧光PCR狐狸 水貂 貉子和狗动物源性成分 

分 类 号：Q819[生物学—生物工程]