多重PCR方法同时鉴别人参、西洋参、三七

目的:建立一种能同时鉴别人参属人参、西洋参、三七3种药材的多重聚合酶链反应(PCR)方法。方法:通过分析人参、西洋参、三七psbA-trnH基因序列的差异设计了特异性引物对1,利用随机扩增多态性DNA(RAPD)分子标记技术筛选获得的特异性片段的基因序列,设计了特异性引物对2;优化了多重PCR鉴别体系并验证其耐受性和实用性。结果:构建的多重PCR鉴别体系能够成功地鉴别人参、西洋参、三七,人参样本可扩增出片段大小约为150 bp的特异性条带,西洋参样本可扩增片段大小出约为150 bp和400bp 2条特异性片段,三七样本可扩增出片段大小约为400 bp的特异性条带。结论:本多重PCR鉴别体系特异性好,可准确、可靠地鉴别人参、西洋参、三七。

Objective: To establish a multiplex polymerase chain reaction(PCR) method for identification of Panax ginseng C.A.Mey., Panax quinquefolium L.and Panax notoginseng F.H.Chen.Methods: The specific primers pair 1 was designed bases on comparison of the difference of the psbA-trnH gene sequences from P.ginseng, P.quinquefolium and P.notoginseng. The pair of specific primers 2 was selected by Random amplified polymorphic DNA(RAPD) molecular marker technology.The multiplex PCR identification system were optimized, and the method's tolerance and suitability was verified.Results: Multiplex PCR identification system for P.ginseng, P.quinquefolium and P.notoginseng was constructed;the of 150 bp specific bands were amplified from the P.ginseng;the sizes of 150 bp and 400 bp specific bands were amplified from P.quinquefolium;and the sizes of 400 bp specific bands were amplified from P.notoginseng.Conclusion: The established multiplex PCR method can be used to identify P.ginseng C.A.Mey., P.quinquefolium L.and P.notoginseng F.H.Chen.

[1] 中国药典2010年版.一部[S].2010:8,131 ChP 2010.Vol I[S].2010:8,131
[2] 中国科学院中国植物志编辑委员会.中国植物志.第五十四卷[M].北京:科学出版社,1999:17 Editorial Committee of flora of China Chinese Academy of Sciences. The Flora of China.Vol 54[M].Beijing:Science Press,1999:17
[3] 佟鹤芳,薛健,童燕玲,等.人参和西洋参特有挥发性成分鉴别[J].中国实验方剂学杂志,2013,21(19):120 TONG HF,XUE J,TONG YL,et al.Analysis of unique volatile components from ginseng and quinquefolius using for determining[J].Chin J Exp Tradit Med Form,2013,21(19):120
[4] 崔秀明,黄璐琦,郭兰萍,等.中国三七产业现状及发展对策[J]. 中国中药杂志,2014,39(4):553 CUI XM,HUANG LQ,GOU LP,et al.Chinese Sanqi industry status and development countermeasures[J].China J Chin Mater Med, 2014,39(4):553
[5] 曹智,张燕娣,许永华,等.人参有效成分及其药理作用研究新进展[J].人参研究,2012,24(2):39 CAO Z,ZHANG YD,XUE YH,et al.Active ingredients and pharmacological activities of ginseng new progress[J].Ginseng Res,2012,24(2):39
[6] 佟鹤芳,薛健,童燕玲,等.人参和西洋参特有挥发性成分鉴别[J].中国实验方剂学杂志,2013,21(19):120 TONG HF,XUE J,TONG YL,et al.Analysis of unique volatile components from ginseng and quinquefolius using for determining[J].Chin J Exp Tradit Med Form,2013,21(19):120
[7] 宋沁馨,冯芳,张心悦,等.SNP测定结合芯片电泳法快速鉴别人参和西洋参[J].药物分析杂志,2009,29(1):1 SONG QX,FENG F,ZHANG XY,et al.Rapid detection the SNP of Panax ginseng and P.quinquefolius by microchip electrophoresis[J].Chin J Pharm Anal,2009,29(1):1
[8] 孙涛,滕少娜,孔德英,等.DNA条形码技术应用于人参鉴定[J].中国生物工程杂志,2013,33(4):143 SUN T,TENG SN,KONG DY,et al.DNA barcoding used in the identification of ginseng[J].China Biotechnol,2013,33(4):143
[9] 詹鑫婕,田程,张媛,等.基于ITS2条形码SNPs的人参和西洋参PCR-SSCP分子鉴别研究[J].中国中药杂志,2012,37(24):3748 ZHAN XJ,TIAN C,ZHANG Y,et al.PCR-SSCP molecular identification of Panax ginseng and P.quinquefolius based on ITS2 barcoding SNPs[J].China J Chin Mater Med,2012,37(24):3748
[10] 曹晖,刘玉萍,伏见裕利,等.三七及其伪品的DNA测序鉴别[J].中药材,2001,24(6):398 CAO H,LIU YP,FUSHIMI H,et al.Panax notoginseng and DNA sequencing of its pseudo products[J].J Chin Med Mater,2001,24(6):398
[11] 田程,刘春生,吴浩忠,等.中药人参与西洋参饮片的电子鼻检测方法及识别模式[J].中国中药杂志,2012,37(8):1165 TIEN C,LIU CS,WU HZ,et al.Detection method and pattern recognition of ginseng and American ginseng pieces by electronic nose[J].China J Chin Mater Med,2012,37(8):1165
[12] 尤超,赵大球,梁乘榜,等.PCR引物设计方法综述[J].现代农业科技,2011,17(3):48 YOU C,ZHAO DQ,LIANG CB,et al.Review of the PCR primer design method[J].Mod Agric Sci Technol,2011,17(3):48