基于ITS2序列的车前子原植物及其混伪品的分子鉴定

目的：通过分析车前子原植物及其混伪品的ITS2条形码序列，探索鉴定车前子及其混伪品的新方法。方法：对研究材料进行DNA提取、PCR扩增和双向测序，所得序列经过CodonCode Aligner拼接后，用软件MEGA5进行相关数据分析，计算其种间、种内遗传距离，并利用已建立的ITS2数据库及其网站预测ITS2二级结构和构建系统发育树。结果：车前种内最大K2P距离为0．0099，与混伪品种间最小K2P距离为0．4976；平车前种内最大K2P距离为0．0052，与混伪品种间最小K2P距离为0．5191。车前子基原植物与其混伪品的二级结构的分子形态均有明显差异。由所构建的系统聚类树可以看出，车前与平车前的不同来源样品聚在一支，并能很好与混伪品区分开。结论：ITS2条形码序列能够成功鉴定车前子基原植物与其混伪品，为车前子的基原鉴定提供了新的方法。

Identification of Original Plant of Plantaginis Semen and Its Adulterants Using DNA Barcoding Method Based on ITS2 Sequence

This study was aimed to explore a new method to identify the original plant of Plantaginis Semen and its adulterants by the ITS2 regions. The second internal transcribed spacer （ITS2） of ribosomal DNA was amplified and sequenced by bidirectional sequencing of PCR products. Sequence assembly and consensus sequence generation were performed by using CodonCode Aligner. The ITS2 secondary structure was predicted using ITS2 database and web- sites. The phylogenetic tree was constructed by MEGA5. The results showed that the maximum intraspecific K2P dis- tance of Plantago asiatica was 0.009 9, while the minimum interspecific K2P distance was 0.497 6; the maximum in- traspeeific K2P distance of P. depressa was 0.005 2, while the minimum interspecific K2P distance was 0.519 1. The ITS2 secondary structure showed that P. asiatica and P. depressa can be differentiated obviously from its adulterants. Different samples of P. asiatica and P. depressa were gathered together and can be distinguished from its adulterants by NJ tree. It was concluded that the ITS2 sequence was able to identify original plant of Plantaginis Semen and its adulterants correctly. It provided a new method for the identification of original plant of Plantaginis Semen.