基于psbK-psbI序列的白鲜鉴定研究  

Identification of Dictamnus dasycarpus Based on psbK-psbI Sequence

作　　者：薛嘉宁 赵容[1] 于莹 高铭泽 尹海波[1] 黄罗强 XUE Jia-ning;ZHAO Rong;YU Ying;GAO Ming-ze;YIN Hai-bo;HUANG Luo-qiang(School of Pharmacy,Liaoning University of Traditional Chinese Medicine,Dalian 116600,China)

机构地区：[1]辽宁中医药大学药学院,辽宁大连116600

出　　处：《中国现代中药》2024年第12期2076-2081,共6页Modern Chinese Medicine

基　　金：辽宁省道地DNA指纹图谱建立项目(2022003)。

摘　　要：目的:基于psbK-psbI序列,运用DNA条形码技术对辽宁省22批与贵州省2批药用植物白鲜的样品进行鉴别分析。方法:利用DNA试剂盒提取法,从24个白鲜样品的根皮中提取DNA,对psbK-psbI序列进行聚合酶链式反应(PCR)扩增后进行双向测序,并从国家生物技术信息中心(NCBI)下载芸香科柑橘属植物香橼和甜橙作为外群序列,运用SeqMan 7.1.0软件对测序结果进行拼接,使用DNAsp 5.1软件统计序列的单倍型数(h)和单倍型多样度(Hd),使用MEGA 7.0软件对数据进行分析比对,计算K2P(Kimura 2-parameter)遗传距离,并利用邻接(NJ)法建立系统发育树进行分析。结果:辽宁省与贵州省共24个白鲜样品的psbK-psbI序列中,21条序列没有变异,3条序列存在1~3个变异位点,结合遗传距离及NJ聚类树的结果可以看出,所有白鲜样品聚为一大支,两种柑橘属的外群聚为一大支,并且支持率为100%,种内差异较小。结论:基于psbK-psbI序列的DNA条形码技术能够对白鲜药材及其同科植物进行区分,证明白鲜这一物种遗传性稳定,可用于白鲜与其他物种进行种间鉴别,为后续白鲜分子水平方面的研究提供理论基础。Objective:To identify Dictamnus dasycarpus from 22 batches of samples from Liaoning province and 2 batches of samples from Guizhou province by DNA barcoding based on the psbK-psbI sequence.Methods:The DNA extraction reagent kit was used to extract DNA from the root bark of 24 D.dasycarpus samples.The psbK-psbI sequence was amplified by polymerase chain reaction(PCR)and then subjected to paired-end sequencing.The sequences of Citrus medica and C.sinensis were downloaded from the National Center of Biotechnology Information(NCBI)as the out-groups.SeqMan 7.1.0 was used to assemble the sequencing results,and DNAsp 5.1 was used to calculate the haplotype number(h)and haplotype diversity(Hd)of the sequences.MEGA 7.0 was used to align the sequences.The Kimura 2-parameter(K2P)model was employed to calculate the genetic distance,on the basis of which the phylogenetic tree was established with the neighbor-joining(NJ)method.Results:Among the psbK-psbI sequences of 24 D.dasycarpus samples from Liaoning and Guizhou,21 sequences showed no variation,while 3 sequences had 1-3 mutation sites.The genetic distance and phylogenetic tree showed that all samples of D.dasycarpus were clustered into a large clade,and the out-groups were clustered into the other clade,with the support rate of 100%,which indicated small intra-species differences.Conclusion:The DNA barcoding based on psbK-psbI sequence can distinguish D.dasycarpus from its homologous plants,which proves the genetic stability of D.dasycarpus.This method is stable and can be used for distinguishing between D.dasycarpu and other species,which provides a theoretical basis for the subsequent research on this plant at the molecular level.

关 键 词：白鲜 psbK-psbI DNA条形码 系统发育分析 

分 类 号：R282[医药卫生—中药学]