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Background: Bupleuri Radix (Chaihu) is a famous traditional Chinese medicine derived from Bupleurum , Apiaceae. The origin of cultivated Chaihu germplasm in China is unclear, which has led to unstable Chaihu quality. In this study, we reconstructed the phylogeny of the main Chaihu germplasm species in China and identified potential molecular markers to authenticate its origin., Methods: Three Bupleurum species (eight individuals), B. bicaule , B. chinense , and B. scorzonerifolium , were selected for genome skimming. Published genomes from B. falcatum and B. marginatum var. stenophyllum were used for comparative analysis., Results: Sequences of the complete plastid genomes were conserved with 113 identical genes ranging from 155,540 to 155,866 bp in length. Phylogenetic reconstruction based on complete plastid genomes resolved intrageneric relationships of the five Bupleurum species with high support. Conflicts between the plastid and nuclear phylogenies were observed, which were mainly ascribed to introgressive hybridization. Comparative analysis showed that noncoding regions of the plastomes had most of the variable sequences. Eight regions ( atp F- atp H, pet N- psb M, rps 16- psb K, pet A- psb J, ndh C- trn V/UAC and ycf 1) had high divergence values in Bupleurum species and could be promising DNA barcodes for Chaihu authentication. A total of seven polymorphic cpSSRs and 438 polymorphic nSSRs were detected across the five Chaihu germplasms. Three photosynthesis-related genes were under positive selection, of which acc D reflected the adaptation fingerprint of B. chinense to different ecological habitats. Our study provides valuable genetic information for phylogenetic investigation, germplasm authentication, and molecular breeding of Chaihu species., Competing Interests: The authors declare there are no competing interests., (©2023 Wang et al.)

Bupleuri Radix is the dry root of certain species of the genus Bupleurum and is commonly used in traditional Chinese medicine. The increasing global demand for Bupleuri Radix cannot be fulfilled with wild populations only. Therefore, cultivated Bupleurum is now the main commercial source of this medicinal product. Different species of Bupleurum show different medicinal properties and clinical effects, making reliable authentication and assignment of correct botanical origin for medicinal species critical. However, accurate identification of the cultivated Bupleurum species is difficult due to dramatic morphological variations resulting from cultivation. In this study, we sampled 56 cultivated Bupleurum populations of six different morphotypes (Types A-F) from the main production areas of China, and 10 wild populations of four species were used as reference materials. Conventional DNA barcoding was conducted to identify cultivated Bupleurum species. Additionally, verification based on complete chloroplast genomes was performed and new chloroplast markers were developed and evaluated. The combination of these methods resulted in the successful identification of all cultivated Bupleurum individuals. Three chloroplast regions are recommended as additional barcodes for the genus: ycf4_cemA, psaJ_rpl33 , and ndhE_ndhG . This is a reliable and promising strategy that can be applied to the authentication of natural products and the identification of other medicinal plant species with similar taxonomic problems., Competing Interests: The authors declare there are no competing interests., (©2022 Zhang et al.)