基于psbA-trnH分子标记的鼠曲草属植物鉴定

Identification of Gnaphalium Herbs based on psbA-trnH molecular markers

目的采用叶绿体基因psbA-trnH间隔区序列对鼠曲草植物、近缘种及其混伪品进行分子鉴定研究,为制定该鼠曲草药材的质量标准提供科学依据,确保其合理使用和进一步的科学发展。方法采集鼠曲草Gnaphalium affine D.Don及其近缘种植物,分别进行DNA基因组提取,经PCR扩增psbA-trnH间隔区序列,进行双向测序,并结合GenBank下载鼠曲草属植物序列,应用MEGA7.0软件分析,进行序列比对、计算遗传距离、构建NJ系统聚类树。结果实验中鼠曲草psbA-trnH序列长度为387~423bp,鼠曲草与其近缘物种psbA-trnH种间平均遗传距离为0.132,种内最大K2P遗传距离为0.005,种间最小K2P遗传距离为0.099,且种间最小遗传距离大于其种内最大遗传距离。基于psbA-trnH序列构建的NJ聚类树显示,鼠曲草属样品聚为一支,其混伪品各自聚为一支。结论psbA-trnH片段可作为鉴别鼠曲草属植物及其易混品种的分子条形码,有利于保障鼠曲草药材的质量安全和临床疗效。

Objective To identify plants of the Gnaphalium,related species and their adulterants by using the chloroplast gene psbA-trnH spacer sequence,so as to provide scientific basis for establishing the quality standard of Gnaphalium,and ensure its rational use and further scientific development.Methods Gnaphalium affine D.Don and its related species plants were collected,genomic DNA were extracted respectively.Then the psbA-trnH spacer sequences were obtained by PCR amplification,and sequenced bidirectionally.The psbA-trnH sequences were aligned together with the sequences of Gnaphalium from GenBank.Then all sequences were analyzed by the software MEGA7.0.The genetic distances were calculated by the Kimura2-parameter(K2 P)model and Neighbor-Joining(NJ)tree constructed with 1000 bootstrap.Results The average interspecific genetic distance among all species was 0.132,the maximum interspecific K2 P genetic distance was 0.005,and the minimum interspecific K2 P genetic distance was 0.099,and the minimum genetic distance between species is greater than the maximum genetic distance within the species.Conclusion The psbA-trnH sequences can be used as a molecular barcode to identify the Gnaphalium plants and their adulterants,which is beneficial to the quality and safety of the Gnaphalium herbs.