岗梅药材及其近缘物种的ITS2分子鉴定研究

Identification of Ilex asprella and Its Closely Related Species Based on ITS2 Barcode

目的 应用ITS2序列对93份岗梅药材及其近缘物种进行鉴定研究。方法 对药材、基原植物样本进行基因组DNA提取,PCR扩增ITS2序列,用MEGA5.0进行序列分析,采用相似性搜索法、最近距离法、构建NJ树对序列鉴定能力进行评估。结果 岗梅药材ITS2序列长度为241 bp,平均GC含量为63.4%,种内K2P遗传距离为0-0.030,岗梅药材及其近缘种的种间最小K2P距离为0.043。岗梅药材和基原物种在NJ树单独聚为一支,能有效地与其他物种分开,相似性搜索法结果表明ITS2序列能够准确鉴定岗梅药材及其近缘种。结论 ITS2序列能够准确鉴定岗梅药材及其近缘物种,可为保障岗梅药材临床用药安全提供新的技术手段。

OBJECTIVE To identify 93 samples of Ilex asprella （ Hook. et Am. ） Champ. ex Benth. and its related species based on ITS2 barcode. METHODS The total genomic DNAs were extracted and the ITS2 regions were amplified and sequenced. The genetic distances were computed using MEGAS. 0 in accordance with the Kimura 2-parameter （K2P） model. The neighbor-joining （NJ） phylogenetic tree and BLAST method were used to assess the efficiency of sequence identification. RESULTS The length of ITS2 sequence of I. asprella was 241 bp and the average GC content was 63.4%. The intra-specific genetic distances of I. asprella were 0 - 0. 030 and the minimum inter-specific of I. asprella and its closely related species was 0. 043. Furthermore, I. asprella formed into a single branch with high bootstrap value in NJ tree. The identification efficiency of ITS2 barcode using BLAST was 100%. CONCLUSION The ITS2 region can accurately and stably distinguish I. asprella from its closely related species and adulterants. It provides a new method to ensure the clinical safety of traditional Chinese medicines.