北柴胡特异DNA 条形码筛选及种质资源鉴定  

Screening of specific DNA barcoding and identification of germplasm resourcesof Bupleurum chinense

作　　者：王馨 尹光耀 张志飞 陈颖 刘珊瑚 满金辉 石玥 黄钰莹 张晓芹[1] 王晓晖[2] 魏胜利[1] WANG Xin;YIN Guang-yao;ZHANG Zhi-fei;CHEN Ying;LIU Shan-hu;MAN Jin-hui;SHI Yue;HUANG Yu-ying;ZHANG Xiao-qin;WANG Xiao-hui;WEI Sheng-li(School of ChineseMateria Medica,Beijing University of Chinese Medicine,Beijing 102488,China;Institute of Chinese Medicine,Beijing University of Chinese Medicine,Beijing 100029,China)

机构地区：[1]北京中医药大学中药学院,北京102488 [2]北京中医药大学中药研究院,北京100029

出　　处：《中草药》2023年第17期5703-5715,共13页Chinese Traditional and Herbal Drugs

基　　金：北京市科学技术委员会基金项目(Z201100005420005);北柴胡精准药材批次分子防伪技术研究项目(2020071720419)。

摘　　要：目的基于北柴胡Bupleurum chinense叶绿体基因组筛选特异性DNA条形码,并利用特异DNA条形码鉴定不同产地北柴胡种质资源。方法利用Illumina HiSeq X Ten平台对北柴胡进行叶绿体基因组测序,利用mVISTA软件和核酸多样性分析筛选特异性片段,基于特异性片段进一步分析不同产区北柴胡样品单倍型。结果不同产地3份北柴胡叶绿体基因组全长为155557~155959 bp,均呈现典型的环状四分体结构,均编码131个基因。trnV-UAC、trnC-GCA_petN、petN_psbM、ndhF_rpl32可以作为潜在的北柴胡种质资源鉴定的特异性DNA条形码。基于扩增效率,选择petN_psbM和ndhF_rpl32对来自4省7个产地177份北柴胡样品进行序列分析,结果表明petN_psbM和ndhF_rpl32分别有91和78个变异位点,分别鉴定到28、29个单倍型;两段序列联合分析形成40个单倍型(Hap1~Hap40),各单倍型的遗传距离为0~0.022。6个产地拥有特有单倍型,可以将不同产地的北柴胡种质资源进行区分。结论利用比较叶绿体基因组学筛选的特异DNA条形码petN_psbM和ndhF_rpl32可以用于北柴胡种内种质资源鉴定,为后续鉴定北柴胡的产地来源、种质资源保护利用和育种等工作奠定基础。Objective To screen the specific DNA barcodes based on Beichaihu(Bupleurum chinense)chloroplast genome,so as to identify the plasm resource of B.chinense from different producing areas.Methods The Illumina HiSeq X Ten platform was used to sequence the chloroplast genome of three samples of B.chinense.Specific DNA barcodes was analyzed with mVISTA soft and nucleotide polymorphism analysis.The haplotype of B.chinense from different producing areas was identified with specific DNA barcodes.Results The total length of chloroplast genomes of three samples from different habitats was 155557—155959 bp,all of which had typical circular tetrad structure and coded 131 genes.trnV-UAC,trnC-GCA_petN,petN_psbM and ndhF_rpl32 can be used as specific DNA barcodes for identification of potential germplasm resources of B.chinense.Based on amplification efficiency,petN_psbM and ndhF_rpl32 were selected to analyse the sequence of 177 samples of B.chinense from seven places in four provinces.The results showed that petN_psbM and ndhF_rpl32 had 91 and 78 mutation loci,and 28 and 29 haplotypes were identified;The two sequences were analyzed together to form 40 haplotypes(Hap1—Hap40),and the genetic distance of each haplotype was 0—0.022.Six producing areas had unique haplotypes,which can distinguish the germplasm resources of B.chinense from different producing areas.Conclusion The specific DNA barcodes petN_psbM and ndhF_rpl32,identified by B.chinense chloroplast genome analysis,could be used to demonstrate the genotypes of B.chinense,which provides values for the identification of B.chinense producing area,the protection and utilization of germplasm resources and breeding work.

关 键 词：北柴胡 叶绿体基因组 序列对比 种质资源 DNA条形码 

分 类 号：R286.12[医药卫生—中药学]