Deciphering the Plastome and Molecular Identities of Six Medicinal "Doukou" Species

Abstract

The genus Amomum includes over 111 species, 6 of which are widely utilized as medicinal plants and have already undergone taxonomic revision. Due to their morphological similarities, the presence of counterfeit and substandard products remains a challenge. Accurate plant identification is, therefore, essential to address these issues. This study utilized 11 newly sequenced samples and extensive NCBI data to perform molecular identification of the six medicinal "Doukou" species. The plastomes of these species exhibited a typical quadripartite structure with a conserved gene content. However, independent variation shifts of the SC/IR boundaries existed between and within species. The comprehensive set of genetic sequences, including ITS, ITS1, ITS2, complete plastomes, matK, rbcL, psbA-trnH, and ycf1, showed varying discrimination of the six "Doukou" species based on both distance and phylogenetic tree methods. Among these, the ITS, ITS1, and complete plastome sequences demonstrated the highest identification success rate (3/6), followed by ycf1 (2/6), and then ITS2, matK, and psbA-trnH (1/6). In contrast, rbcL failed to identify any species. This research established a basis for a reliable molecular identification method for medicinal "Doukou" plants to protect wild plant resources, promote the sustainable use of medicinal plants, and restrict the exploitation of these resources.

Keywords: DNA barcoding; ITS; medicinal plants; plastome; species identification; “Doukou”.

Figure 1

Plastome gene map of Wurfbainia compacta YWB91902-2 showing the typical structure organization in “Doukou” plastomes. Genes inside the circle are transcribed clockwise, and those outside are transcribed counterclockwise. Genes in different functional groups are color-coded. The small and large single-copy regions (SSC and LSC) and inverted repeat (IRa and IRb) regions are noted in the inner circle.

Figure 2

Comparison of the borders of the LSC, SSC, and IR regions among six plastomes of Amomum. Abbreviations: JLB—Junction of large single-copy and small single-copy regions; JSB—Junction of small single copy and inverted repeat B; JSA—Junction of small single copy and inverted repeat A; JLA—Junction of large single copy and inverted repeat A.

Figure 3

The variable sites in the homologous regions of 41 plastomes of “Doukou” species and their taxonomic synonyms. The y-axis represents the nucleotide diversity (Pi), and the x-axis indicates the nucleotide midpoints.

Figure 4

Scatter plot of barcoding gap analysis of the eight datasets across the six medicinal “Doukou” species and their taxonomic synonyms. The y-axis represents the genetic divergence, with the plots above the blue line of best fit representing successfully delimited species and those along and below the line representing the overlap. “CP” represents complete plastome.

Figure 5

The phylogenetic tree was reconstructed using the maximum likelihood (ML) method with the ITS dataset of six medicinal “Doukou” species and their taxonomic synonyms. The numbers at nodes indicate bootstrap values.

Figure 6

The phylogenetic tree was reconstructed using the maximum likelihood (ML) method with the complete plastome dataset of six medicinal “Doukou” species and their taxonomic synonyms. The numbers at nodes indicate bootstrap values.

Figure 7

The phylogenetic tree was reconstructed using the maximum likelihood (ML) method with the ITS1 dataset of six medicinal “Doukou” species and their taxonomic synonyms. The numbers at nodes indicate bootstrap values.

Figure 8

The species discrimination success for candidate barcodes of six medicinal “Doukou” plants across different delimitation methods. The success rate is the number of species successfully delimited to species in the different DNA markers. “CP” represents complete plastome.