北苍术特异性DNA条形码筛选、种质资源鉴定及遗传多样性分析  

Screening of specific DNA barcode, identification of germplasm resources, and analysis of genetic diversity of Atractylodes chinenesis

作　　者：尹光耀 袁林 王馨 张志飞 陈颖 刘珊瑚 满金辉 石玥 黄钰莹 张晓芹[1] 王晓晖 魏胜利[1,4] YIN Guang-yao;YUAN Lin;WANG Xin;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 Chinese Materia Medica,Beijing University of Chinese Medicine,Beijing 102488,China;National Medical Products Administration,Beijing 100037,China;Modern Research Center for Traditional Chinese Medicine,Beijing Institute of Traditional Chinese Medicine,Beijing University of Chinese Medicine,Beijing 102488,China;Engineering Research Center of Good Agricultural Practice for Chinese Crude Drugs of Ministry of Education,Beijing 100102,China)

机构地区：[1]北京中医药大学中药学院,北京102488 [2]国家药品监督管理局,北京100037 [3]北京中医药大学北京中医药研究院中药现代研究中心,北京102488 [4]中药材规范化生产教育部工程研究中心,北京100102

出　　处：《药学学报》2023年第6期1693-1704,共12页Acta Pharmaceutica Sinica

基　　金：北苍术、赤芍精准药材批次分子防伪技术研究项目(2020110037009381);北京市科学技术委员会基金项目(Z201100005420005)。

摘　　要：北苍术Atractylodes chinensis具有重要药用价值和经济价值。本研究利用Illumina平台测序获得4份不同产地的北苍术的叶绿体全基因组序列,筛选特异DNA条形码,并利用特异DNA条形码对不同产区北苍术样品种质资源鉴定及居群的遗传多样性分析。该4份北苍术叶绿体全基因组均为典型的环状四分体结构,均注释112个基因。比较基因组学研究结果表明ccsA和trnC-GCA_petN是潜在的北苍术种内鉴别的特异DNA条形码。选择ccsA和trnC-GCA_petN对来自9省14产区的256份样品进行PCR扩增,扩增效率为100%。序列分析结果表明ccsA和trnC-GCA_petN分别有11和22个变异位点,分别能鉴定16和22个单倍型,两段序列联合分析鉴定39个单倍型,命名为Hap1~Hap39,其中占比最多和分布最广的基因型为Hap9。单倍型多样性(Hd)=0.896,核苷酸多样性(Pi)=0.002 22,说明北苍术在物种水平上有较高的遗传多样性。各单倍型的遗传距离为0.000 00~0.004 88,说明各个单倍型之间有较小的遗传差异。系统进化树分析结果表明, 39个单倍型有很近的亲缘关系,并且与除白术外其余同属类群形成明显的两个分支。本研究为后续鉴定北苍术产地来源和后续种质资源保护和育种方面起到重要作用。Atractylodes chinensis has important medicinal and economic values.In this study,the chloroplast genome sequences of four A.chinensis samples from different producing areas were sequenced using the Illumina platform.The specific DNA barcodes were screened and the germplasm resources of A.chinensis samples from different producing areas and the genetic diversity of the population were analyzed basing on the specific barcodes.The whole chloroplast genomes of the four A.chinensis samples had a typical cyclic tetrad structure,with 112 genes annotated.The comparative genomics results indicated that ccsA and trnC-GCA_petN were potential specific DNA barcodes for intraspecific identification of A.chinensis.Polymerase chain reaction(PCR)analysis of ccsA and trnC-GCA_petN was performed on 256 samples from 14 areas in 9 provinces,and the amplification efficiency was 100%.Sequence analysis showed that ccsA and trnC-GCA_petN had 11 and 22 variant positions,which could identify 16 and 22 haplotypes,respectively.The combined sequence analysis identified 39 haplotypes,named Hap1-Hap39,of which the most abundant and widely distributed genotype was Hap9.Haplotype diversity(Hd)=0.896 and nucleotide diversity(Pi)=0.00222 indicated high genetic diversity at the species level in A.chinensis.The genetic distances of the haplotypes were 0.00000-0.00488,indicating that there were small genetic differences among the haplotypes.The results of phylogenetic tree analysis showed that 39 haplotypes had very close genetic relationship,and formed two obvious branches with other groups of the same genus except Atractylodes macrocephala.This study plays an important role in the identification of the origin of A.chinensis and the protection and breeding of germplasm resources.

关 键 词：苍术 叶绿体基因组 DNA条形码 单倍型 遗传多样性 

分 类 号：R931[医药卫生—生药学]