甘肃中医药大学学报2026，Vol.43Issue(2)：38-44,7.DOI:10.16841/j.issn1003-8450.2026.02.08
基于DNA条形码的紫花地丁、苦地丁及甜地丁鉴定
Authentication of Viola philippica,Corydalis bungeana and Gueldenstaedtia verna based on DNA barcoding
摘要
Abstract
Objective To authenticate Viola philippica(Zihuadiding),Corydalis bungeana(Kudiding),and Gueldenstaedtia verna(Tiandiding)using the nucleotide sequence of the second internal transcribed spacer(ITS2),a DNA barcode marker.Methods Total genomic DNA was extracted from samples via a column-based plant ge-nomic DNA extraction kit.Fluorescent quantitative polymerase chain reaction(PCR)amplification was performed using universal ITS2 primers;following optimization of the annealing temperature,bidirectional sequencing was con-ducted on target bands.The obtained sequences were assembled with CodonCode Aligner 11.0.1,and primer and flanking sequences were trimmed.Genetic distances and parsimony-informative sites were calculated using MEGA 11.0 software,and a neighbor-joining(NJ)phylogenetic tree was constructed.Additionally,the secondary structure of the ITS2 sequence was predicted online.Results The intraspecific genetic distances of V.philippica,C.bun-geana,and G.verna were 0.01,0.00,and 0.00,respectively,while interspecific genetic distances ranged from 0.69 to 0.82.Specifically,the genetic distance(0.69)and number of parsimony-informative sites(93)between V.philippica and C.bungeana were lower than those between V.philippica and G.verna(0.82,102).The interspecific genetic dis-tance between G.verna and C.bungeana was 0.81,with 100 parsimony-informative sites.The NJ phylogenetic tree showed that each species formed a monophyletic clade;V.philippica and C.bungeana clustered into the same major branch,which was sister to the clade of G.verna.Analysis of the ITS2 secondary structure revealed that V.philippica lacked the typical"one-ring and four-arms"configuration,whereas both C.bungeana and G.verna possessed this structure—with distinct differences in the apical loop of arm Ⅲ and the middle loop of arm Ⅳ between the latter two.Conclusion ITS2 sequences combined with their secondary structures can effectively distinguish V.philippica,C.bungeana,and G.verna,and clarify their phylogenetic relationships:V.philippica and C.bungeana are closely rela-ted.This method provides a reliable and reproducible molecular basis for the accurate identification of Dididing-class medicinal materials.关键词
紫花地丁/苦地丁/甜地丁/DNA条形码/邻接系统发育树/二级结构Key words
Viola philippica/Corydalis bungeana/Gueldenstaedtia verna/DNA barcoding/neighbor-joining phy-logenetic tree/secondary structure分类
医药卫生引用本文复制引用
王孟虎,金阳,孟祥松,栗进才,俞浩,牛倩,孙一帆..基于DNA条形码的紫花地丁、苦地丁及甜地丁鉴定[J].甘肃中医药大学学报,2026,43(2):38-44,7.基金项目
安徽省教育厅高校科学研究项目(2023AH053216) （2023AH053216）
安徽省教育厅优秀青年教师培育一般项目(YQYB2024085) （YQYB2024085）
安徽高校协同创新项目(GXXT-2023-073) （GXXT-2023-073）
亳州学院质量工程项目(2024XJXM041). （2024XJXM041）
