Classification and authentication of six Taraxacum species based on complete chloroplast genomes, ribosomal DNAs, and metabolite profiling

Abstract

민들레속 식물들은 오랫동안 전통 한약재로서 사용되어 왔다. 각 식물체의 건조된 상태는 구분하기가 매우 어려워 한약재나 제품으로 사용될 때 혼용되는 경우가 빈번히 발생하고 있는 실정이다. 이에 민들레 종들을 체계적으로 분류할 수 있는 바코드 마커를 개발하기 위해 엽록체와 45S nuclear ribosomal DNA 서열을 분석하였다. NGS 기술을 이용하여 대한민국 공정서에 포공영으로 등록된 4종 (서양민들레, 민들레, 털민들레, 흰민들레)와 근연식물(산민들레, 붉은씨서양민들레)에 대해 엽록체와 핵 내 ITS 영역을 포함하는 45S rDNA 전체서열을 완성하였다. 완성된 서열의 변이지역들을 기반으로 공정서 등록종과 근연식물을 비교하여 포공영 등록종을 안정적으로 판별할 수 있는 바코드 마커 시스템을 구축하였다. 또한 민들레속 식물들에 대해 대사체 비교 분석을 수행하였으며, 각각의 종 특이 대사체 마커를 선정하였다. 본 연구에서 획득한 완전한 엽록체 genome과 45S nrDNA 서열에서 밝혀진 다양한 종간 변이는 민들레속 식물의 근연관계 확인 및 국화과 근연종 다양성 연구에 가치 있게 활용될 수 있을 것이다. 그리고 본 연구에서 개발한 바코드 마커는 시장에서 유통되는 민들레 유래 약재 및 제품을 판별하기 위해서 사용될 수 있을 것이다.

Taraxacum species, known as dandelion, are perennial plants in the family Asteraceae, the largest family of flowering plants. The genus Taraxacum consists of over 2,500 species which is widely distributed in the warm temperate areas of the Northern Hemisphere. Taraxacum species have long been utilized for medicinal purposes due to their various pharmacological activities including diuretic and anti-inflammatory effects. In Korean official compendium for herbal medicine, Po gong young is dried entire part of T. officinale, T. platycarpum, T. mongolicum, and T. coreanum. The genus Taraxacum shows taxonomic complexity because of the presence of agamospermy, complex hybrids, and polyploidy. Six Taraxacum species have different morphology, but it is hard to distinguish these plants in dried or processed state. For authentication of these six species and development of barcode markers, chloroplast (cp) genome and 45S nuclear ribosomal DNA (45S nrDNA) sequences were analyzed. Complete cp genome and 45S nrDNA sequences were characterized using the low coverage of whole genome sequences produced by next-generation sequencing technology. The complete cp genomes of the six Taraxacum species range from 151,173 bp to 151,451 bp. All six cp genomes share the typical quadripartite chloroplast structure of angiosperm. The 45S nrDNA units of six Taraxacum species were 5,826 bp in length. The six Taraxacum chloroplast genomes identically harbored a total of 112 genes arranged in the same order, including 79 protein-coding genes, 29 transfer RNA genes, and four ribosomal RNA genes. Divergent sequence variations of SNPs and InDels were identified among six Taraxacum species. A total of 756 SNPs and 133 InDels in cp genomes and 71 SNPs were identified in 45S rDNA genomes among six Taraxacum species. The phylogenetic relationships based on chloroplast and 45S nrDNA sequences revealed that registered species of Po gong young and not-registered species were clustered together. 53 Tandem repeats longer than 10 bp were detected in cp genomes of six Taraxacum species. Three InDel-based markers derived from cp sequences were developed and successfully validated to authenticate six species and commercial samples of Po gong young. Metabolite profiles of five species Taraxacum species were analyzed, distinct metabolite patterns of each species were identified. Integrative analysis of genomics and metabolite profiling suggests that reliable taxonomy systems of different Taraxacum species.