多重位点特异性PCR鉴别人参、三七、西洋参掺杂-2019年文档.docVIP

多重位点特异性PCR鉴别人参、三七、西洋参掺杂 　　Identification of Panax ginseng， P. notoginseng and P. quinquefolius 　　admixture by multiplex allelespecific polymerase chain reaction 　　JIANG Chao， LUO Yuqing， YUAN Yuan*， HUANG Luqi， JIN Yan， ZHAO Yuyang 　　（State Key Laboratory of Daodi Herbs Breeding Base， National Resource Center for Chinese 　　Materia Medica， China Academy of Chinese Medical Sciences， Beijing 100700， China） 　　[] To achieve a molecular method to identify Panax ginseng， P. notoginseng，P. quinquefolius and their admixture. The ITS，18S and matK sequences of Panax genus were analyzed to develop speciesspecific SNP marker. Three pairs of speciesspecific primers were designed to establish a multiplex allelespecific polymerase chain reaction （MASPCR） and the samples from different regionwere tested. The results showed that when the annealing temperature was 60 ℃ and the cycle number was 35， approximately 250， 500，1 000 bp specific band were obtained from P. ginseng， P. notoginseng and P. quinquefolius obtain， respectively. This method could also be used to authentificate admixture samples and could detect 0.5% percent of P. notoginseng or P. quinquefolius adulterated in P. ginseng， or 0.5% percent of P. ginseng or P. quinquefolius adulterated in P. notoginseng. The detect limit of P. ginseng in P. quinquefolius was 0.5% and P. notoginseng in P. quinquefolius was 1%. This results showed that the present method could be used as a promise method to identify Panax ginseng， P. notoginseng， P. quinquefoliusand their admixture. 　　人、西洋参、三七是我国常用名贵中药，其根、根茎、叶、花、果实等均做药用，价值很高。在中医临床用药上，人参性温，功效大补元气、固脱生津、安神；三七性温，功效散瘀止血、消肿定痛；西洋参性凉，功效补气养阴、清热生津，三者药效具有很大区别，不能混用。然而，由于人参、三七、西洋参为人参属近缘物种，形态和化学成分相似，市场时有掺杂混用现象发生，人参、西洋参饮片或人参花、西洋参花、三七花相互掺杂尤为常见，对用药安全造成损害，需要建立准确、可靠、特别是能同时检测是否存在相互掺杂的鉴定方法。 　　分子鉴定尤其是基于SNP的方法，因其具有准确性高、特异性好、不受环境影响等优点，在人参属物种鉴定中已有应用，如聚合酶链式反应单链构象多态性（PCRSSCP）[1]、聚合酶链式反应限制性内切酶酶切图谱多态性（PCRRFLP）[2]、位点特异性PCR或多重PCR[36]以及序列测定技术[6]等。但这些技术只关注人参属真伪品的鉴定，无法鉴定是否存在掺杂品，对市场实际存在的花、叶、饮片类掺杂样品需要进行多次PCR反应才能判定。本研