Comparative chloroplast genome analyses provide new insights into molecular markers for distinguishing Arnebiae Radix and its substitutes (tribe Lithospermeae, Boraginaceae)

Abstract

Background: Arnebiae Radix has long been used in traditional medicine for its pleiotropic properties. However, distinguishing Arnebiae Radix from its substitutes or closely related species has been challenging due to limited phenotypic characteristics.

Purpose: We aimed to identify the molecular markers for distinguishing Arnebiae Radix from its confusion species.

Study design: Chloroplast genome sequences were used to identify the markers.

Methods: Chloroplast genomes from 15 species across five genera, including all historically used source plants, were sequenced and assembled.

Results: The results revealed significant similarities across all chloroplast genomes in terms of structure, size, gene content, repeat sequences pattens, and codon usage patterns. Phylogenetic analysis showed that the genera Lithospermum, Buglossoides, and Aegonychon formed one clade, while Arnebia guttata, Ar. decumbens, and Ar. euchroma form another. Despite most regions of the chloroplast genomes are highly conserved, three regions-petA-psbJ, ndhF-rpl32, and ycf1-exhibited high variability among difference species, providing high-resolution markers for species identification. Specifically, 376 and 325 species-specific sites were identified in Ar. euchroma and Ar. guttata, respectively. Additionally, four species-specific sites were identified as novel molecular markers, potentially aiding in distinguishing Arnebiae Radix and its confusion or substitute species.

Conclusion: This study provided new genetic insights for differentiating Arnebiae Radix and its confusion species, paving the way for further exploration of these medicinal plants.

Keywords: Arnebiae radix; Chloroplast genome; Genetic divergence; SNPs; Species identification.