基于DNA条形码技术鉴定蒙药材刺柏叶及其混淆品

Identification of Juniperus Folium the Mongolian Medicine and Its Adulterants Based on DNA Barcoding Technique

目的:基于DNA条形码技术鉴别蒙药材刺柏叶及其混淆品。方法:采用国际通用的条形码序列ITS2、psb A-trn H、mat K和rbc L,对刺柏叶及其混淆品圆柏叶共计11份样品进行DNA提取、扩增,采用CodonCode Aligner进行序列拼接,并用MEGA软件对拼接序列进行变异位点分析、邻接(NJ)聚类分析,并计算其平均种内、种间遗传距离。结果:4对引物的PCR扩增产物测序成功率分别为ITS2 100%、psb A-trn H 100%、rbc L 100%、mat K 0%;ITS2序列和psb A-trn H序列均可通过变异位点比较区分刺柏叶及其混淆品;NJ聚类分析的结果显示,psb A-trn H序列的NJ聚类树中刺柏叶与圆柏叶均能分别聚为一支,且psb A-trn H序列的平均种间遗传距离明显大于平均种内遗传距离。结论:psb A-trn H序列能有效区分圆柏叶与刺柏叶,可作为鉴别刺柏叶及其混淆品圆柏叶的条形码序列,为蒙药材刺柏叶及其混淆品的鉴别提供支持。

Objective:To identify the Juniperus Folium of Mongolian medicine and its adulterants by DNA barcode technology.Method:The DNA extraction and amplification of 11 samples of Juniperus Folium and the adulterants Sabina Folium were carried out by using the internationally recognized barcode sequences ITS2,psbA-trnH,matK and rbcL.Codon Code Aligner was adopted to align the sequence,and MEGA software was used to analyze the mutation sites and Neighbor-Joining(NJ)cluster of the splice sequence,and its average intra-specific and inter-specific genetic distance were calculated.Results:The success rates of PCR amplification products of the four primers were ITS2100%,psbA-trnH 100%,rbcL 100%and matK 0%,respectively.The ITS2 sequences and psbA-trnH sequences could distinguish the Juniperus Folium and its adulterants by comparison of variation sites.The results of NJ cluster analysis showed that the psbA-trnH sequences of Juniperus Folium and its adulterants could be clustered into one branch respectively,and the average interspecific genetic distance of psbA-trnH sequences was significantly greater than the average intraspecific genetic distance.Conclusion:The psbAtrnH sequences can effectively distinguish Juniperus Folium from Sabina Folium,and can be used as barcode sequence to identify Juniperus Folium and the adulterants Sabina Folium,which will provide support for the identification of Mongolian medicinal materials Juniperus Folium and the adulterants Sabina Folium.