药用植物三尖杉及其同属易混物种的DNA条形码鉴定研究

Studies on DNA Barcoding Identification for Cephalotaxus fortunei and Its Common Species in the Same Genus

目的:探索建立适用于鉴定药用植物三尖杉及其同属易混物种的DNA条形码鉴定体系,以保障其用药安全。方法:提取三尖杉属3个物种(三尖杉、粗榧和篦子三尖杉)共22份样本的基因组DNA、扩增ITS2和psb A-trn H序列并测序,通过Codon Code Aligner V4.2对测序峰图进行校对拼接。应用MEGA 5.1计算种内和种间Kimura 2-Paramter(K2P)遗传距离,构建邻接(Neighbor-Joining,NJ)系统聚类树。结果:基于ITS2序列分析,三尖杉的种内最大遗传距离小于种间最小遗传距离;基于psb A-trn H序列分析,三尖杉的种内和种间遗传距离有重合。结论:应用ITS2序列,可以实现对三尖杉及其易混物种的鉴别,为三尖杉药材的安全使用提供鉴定依据。

This study was aimed to explore the establishment of DNA barcode identification system for the identification of Cephalotaxus fortunei and its common species in the same genus, which was significance to the safety of drug use. In this paper, 22 samples from 3 species（C. fortunei, C. sinensis and C. oliveri） in genus Cephalotaxus were studied, whose genomic DNA was extracted and amplified by PCR to obtain both ITS2 and psb A-trn H sequences. Then, Codon Code Aligner V4.2 was used to splice sequences, and MEGA 5.1 was used to calculate the intraspecific and interspecific Kimura 2-Parameter（K2 P） genetic distances in order to build Neighbor-Joining（NJ） phylogenetic trees. The results showed that based on ITS2 sequences, the maximum K2 P intraspecific genetic distance of C. fortunei was lower than the minimum K2 P interspecific genetic distance among it and its adulterants. While based on psb A-trn H sequences, the genetic distance between the intraspecific and interspecific of C. fortunei had a coincidence. It was concluded that ITS2 sequences can be used for identification of C. fortunei and its common species in the same genus, which will provide an identification basis for the safe use of C. fortunei.