Molecular Identification of Isatidis Folium and Its Adulterants by ITS2 Sequences
大青叶及其混淆品的ITS2序列鉴定研究

This study aimed to discriminate between Isatidis Folium and its adulterants in order to guarantee the quality and clinical curative effect of this medical material.In this study,the second internal transcribed spacer(ITS2) of ribosomal DNA were amplified and sequenced.Sequence assembly and consensus sequence generation were performed by using the CodonCode Aligner.Phylogenetic study was performed by using software MEGA 4.0 software in accordance with the Kimura 2-parameter(K2P) model.And the phylogenetic tree was constructed using the neighbor-joining methods.The ITS2 secondary structure was predicted by using the ITS2 database and website which was built by Koetschan et al.The results showed that the length of ITS2 sequence of the origin plant of Isatidis Folium was 191 bp.The mean intraspecific genetic distance(K2P distance,0.002) was far lower than its mean interspecific genetic distance(0.882).In the cluster dendrogram,all species showed monophyletic,and distinguished from others.To compare the ITS2 secondary structure of the origin plant of Isatidis Folium and its adulterants,we noticed that it was obviously distinguished from other species in the amount,size,position of loop and the angle of helix exertion.It is concluded that ITS2 sequence can be used as DNA barcode to identify Isatidis Folium and its adulterants effectively.This study may provide an important example for the authentication of other medicinal herbs listed in the Chinese Pharmacopoeia.