中药辛夷及其近缘种的DNA条形码分子鉴定  

Molecular Identification of Magnoliae Flos and Its Relatives Based on DNA Barcoding

作　　者：张心怡 邱雨轩 胡杨 田荣[2] 赵勉 严辉 王吓长[1,2] 胡立宏 ZHANG Xin-yi;QIU Yu-xuan;HU Yang;TIAN Rong;ZHAO Mian;YAN Hui;WANG Xia-chang;HU Li-hong(Jiangsu Key Laboratory for Functional Substances of Chinese Medicine,Nanjing University of Chinese Medicine,Nanjing 210023,China;School of Pharmacy,Nanjing University of Chinese Medicine,Nanjing 210023,China)

机构地区：[1]南京中医药大学江苏省中药功效物质重点实验室,江苏南京210023 [2]南京中医药大学药学院,江苏南京210023

出　　处：《南京中医药大学学报》2023年第11期1086-1094,共9页Journal of Nanjing University of Traditional Chinese Medicine

基　　金：国家自然科学基金青年科学基金项目(81803391);江苏省高等学校基础科学(自然科学)研究重大项目(23KJA350003);江苏省研究生实践创新计划项目(KYCX22_2041);江苏高校“青蓝工程”资助项目。

摘　　要：目的 应用DNA条形码技术对中药辛夷及其近缘种进行分子鉴定,建立快速、准确、便捷的辛夷药材鉴定方法。方法 提取21份辛夷药材基原植物及其近缘种基因组DNA,扩增ITS、ITS2、psbA-trnH、matK和rbcL基因序列并进行双向测序;运用MEGA7.0软件进行序列比对,基于Kimura-2-Parameter(K2P)模型计算种内、种间的遗传距离以评估Barcoding gap,构建邻接(NJ)系统发育树反应鉴定结果。结果 5个引物序列中,psbA-trnH和matK序列的2个引物能正常扩增目的基因,测序成功率均大于98%,rbcL测序成功率较低,只有38.1%;其中matK具有最多的变异位点,psbA-trnH较matK和rbcL的Barcoding gap明显;NJ系统发育树显示,psbA-trnH+matK组合条形码序列能够准确鉴别9个品种,将辛夷药材及其近缘种很好地区分,并首次将其应用到玉兰嫁接砧木品种的分子鉴定。结论 应用psbA-trnH+matK的序列组合能够实现辛夷药材的准确鉴定,并且可以应用于玉兰嫁接砧木品种的快速鉴别。OBJECTIVE To identify Magnoliae Flos and its related species using DNA barcoding technology,so as to establish a rapid,accurate and convenient method for identification of Magnoliae Flos.METHODS Genomic DNA of 21 Magnoliae Flos and their related species were extracted,and ITS,ITS2,psb A-trn H,mat K and rbc L gene sequences were amplified and sequenced.MEGA7.0 software was used for sequence comparison,and the intraspecific and interspecific genetic distance was calculated based on Kimura-2-Parameter(K2P)model to evaluate Barcoding gap,and the NJ phylogenetic trees were constructed to react identification results.RESULTS Among the 5 primers,psb A-trn H and mat K could normally amplify the target gene,and the success rate of them was higher than 98%,while the success rate of rbc L sequencing was 38.1%only.The mat K sequence had the highest number of variation sites,and psb A-trn H had the most prominent species discriminatory on Barcoding gap analysis relative to mat K and rbc L.The NJ phylogenetic tree showed that psb A-trn H+mat K barcode sequence could accurately identify 9 varieties,and distinguish Magnoliae Flos and its relatives.In addition,we applied it to the molecular identification of Magnolia grafting rootstocks for the first time.CONCLUSION The sequence combination of psb A-trn H+mat K can realize the accurate identification of Magnoliae Flos,and can be applied to the rapid identification of Magnolia grafting rootstocks.

关 键 词：辛夷 DNA条形码 分子鉴定 psbA-trnH+matK 嫁接繁殖 

分 类 号：R282.5[医药卫生—中药学]