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[摘要]
目的 比较研究野豌豆属中入蒙药的8种药用植物山野豌豆Vicia amoena、北野豌豆V. ramuliflora、广布野豌豆V. cracca、多茎野豌豆V. multicaulis、黑龙江野豌豆V. amurensis、东方野豌豆V. japonica、大叶野豌豆V. pseudorobus和歪头菜V. unijuga的rDNA-ITS序列差异性及其规律, 寻找8种植物药之间的分子鉴别方法。方法 从野豌豆属蒙药材中提取总DNA, 用PCR法扩增ITS, 运用Clustal X、MEGA等软件比较和分析ITS的序列特征。结果 野豌豆属入蒙药的8种药用植物ITS序列长度为598~611 bp, 具变异位点40个, 其中信息位点33个, 特异性鉴别位点29个。序列已提交至NCBI数据库, 登录号为JQ309787~JQ309794, KJ417905~KJ417931。其中5种野豌豆的ITS序列为国际首次获得, 分别为黑龙江野豌豆(JQ309789)、东方野豌豆(JQ309792)、多茎野豌豆(JQ309791)、大叶野豌豆(JQ309788)和北野豌豆(JQ309793)。结论 ITS序列对野豌豆属入蒙药的8种药用植物具有较好的分辨性, 为野豌豆属蒙药药用植物分子鉴别提供了参考。
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[Abstract]
Objective To find the patterns of the rDNA ITS sequence variation of eight traditional Mongolian medicinal plants in Vicia L. (including V. amoena, V. ramuliflora, V. cracca, V. multicaulis, V. amurensis, V. japonica, V. pseudorobus, and V. unijuga), and establish the molecular biological method for the identification of the eight plants. Methods DNA was extracted from the dry leaves, and the ITS gene fragments were PCR amplified and sequenced. The sequence features were compared and analyzed using Clustal X and MEGA softwares. Results The length of ITS region varied from 598—611 bp with 40 variable sites, 33 parsimony information sites, and 29 singleton sites among eight traditional mongolian medicinal plants in Vicia L. Sequences were submitted to NCBI database with the registry numbers of JQ309787—JQ309794, KJ417905—KJ417931. And ITS sequences of five species among them were first reported in the world, they were V. amurensis (JQ309789), V. japonica (JQ309792), V. multicaulis (JQ309791), V. pseudorobus (JQ309788), and V. ramuliflora (JQ309793). Conclusion ITS sequences could be used to authenticate the eight species in Vicia L. and also could provide a reference for molecular identification.
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[基金项目]
内蒙古科技厅科技计划项目(20080502);内蒙古医科大学科技百万工程项目(kjbw2012009)