title: Development of molecular markers, based on chloroplast and ribosomal DNA regions, to discriminate three popular medicinal plant species, Cynanchum wilfordii, Cynanchum auriculatum, and Polygonum multiflorum.
abstract: Identification of plant species is important for standardizing herbal medicine. Cynanchum wilfordii (Baekshuoh in Korean) and Polygonum multiflorum (Hashuoh in Korean) are important oriental medicinal herbs in Korea, Japan, and China. Cynanchum auriculatum is a faster growing and more productive plant than C. wilfordii; and, it is not recognized as a medicinal plant in the Korean Pharmacopoeia. C. wilfordii, P. multiflorum, and C. auriculatum are often misidentified in the Korean herbal medicine marketplace due to their morphological similarities and similar names. In this study, we investigated molecular authentication of these three medicinal plants using DNA sequences in the TrnL-F chloroplast intergenic region. Specific species identification was achieved by detecting allelic variations of single nucleotide polymorphisms (SNPs) using amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) and high resolution melting curve analysis. Our results demonstrate that the intraspecific genetic distance between C. wilfordii and C. auriculatum is relatively low. We also developed a quantitative PCR assay using species-specific TrnL-F primers, which allowed us to estimate the ratio of C. wilfordii and C. auriculatum using varying ratios of mixed genomic DNA template from the two species. Additionally, to identify species in hybrid plants produced by cross-fertilization, we analyzed nuclear ribosomal DNA internal transcribed spacer regions in C. wilfordii and C. auriculatum by ARMS-PCR. Our results indicate that SNP-based molecular markers, usable to barcode tools could provide efficient and rapid authentication of these closely related medicinal plant species, and will be useful for preventing the distribution of products contaminated with adulterants.
authors.affiliation: Department of Agonomy & Medicinal Plant Resources, Gyeongnam National University of Science & Technology, JinJu, 660-758, Republic of Korea.
authors.forename: Eun-Heui
authors.initials: EH
authors.lastname: Han
authors.affiliation: Department of Food Science, Gyeongnam National University of Science & Technology, JinJu, Republic of Korea.
authors.forename: KyeMan
authors.initials: K
authors.lastname: Cho
authors.affiliation: Gyeongnam Oriental Medicinal Herb Institute, Sancheong, Republic of Korea.
authors.forename: YoungMin
authors.initials: Y
authors.lastname: Goo
authors.affiliation: Gyeongsangnam-do Agricultural Research & Extension Services, JinJu, 660-985, South Korea.
authors.forename: ManBae
authors.initials: M
authors.lastname: Kim
authors.affiliation: Department of Agonomy & Medicinal Plant Resources, Gyeongnam National University of Science & Technology, JinJu, 660-758, Republic of Korea.
authors.forename: Young-Wook
authors.initials: YW
authors.lastname: Shin
authors.affiliation: Department of Biology Education, College of Education, and IALS, Gyeongsang National University, JinJu, 660-701, Republic of Korea. cefle@gnu.ac.kr.
authors.forename: Yun-Hee
authors.initials: YH
authors.lastname: Kim
authors.affiliation: Department of Agonomy & Medicinal Plant Resources, Gyeongnam National University of Science & Technology, JinJu, 660-758, Republic of Korea. shinwlee@gntech.ac.kr.
authors.forename: Shin-Woo
authors.initials: SW
authors.lastname: Lee
mesh_terms: D001483:Base Sequence; D031105:Cynanchum / Q000145:classification / Q000235:genetics*; D058893:DNA Barcoding, Taxonomic; D018742:DNA, Chloroplast / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics*; D021903:DNA, Ribosomal Spacer / Q000235:genetics*; D000068676:Fallopia multiflora / Q000145:classification / Q000235:genetics*; D008969:Molecular Sequence Data; D010946:Plants, Medicinal / Q000145:classification / Q000235:genetics; D020641:Polymorphism, Single Nucleotide; D060888:Real-Time Polymerase Chain Reaction; D016415:Sequence Alignment
chemical_list: D018742:DNA, Chloroplast; D018744:DNA, Plant; D021903:DNA, Ribosomal Spacer
keywords: ARMS-PCR; Chloroplast genome; HRM curve analysis; Nuclear ribosomal DNA internal transcribed spacers; Single nucleotide polymorphisms