The complete chloroplast genome sequences of monotypic genus Pseudogalium, and comparative analyses with its relative genera

Abstract

Background: Pseudogalium is a new monotypic genus with two subspecies in China and one in Japan, which holds a distinctive phylogenetic position and ecological significance within the tribe Rubieae. Chloroplast genomes contain abundant information for resolving phylogenetic relationships. To investigate the phylogenetics of P. paradoxum and its related genera, we first sequenced, assembled, and annotated the chloroplast genome of two subspecies of P. paradoxum in China and reconstructed the phylogenetic trees. Due to the lack of samples of P. paradoxum subsp. franchetianum from Japan, this study only analyzed and discussed P. paradoxum subsp. paradoxum and P. paradoxum subsp. duthiei.

Results: This study had shown that the complete chloroplast genomes of Pseudogalium ranged from 153,093 bp to 153,333 bp in length with 130 genes in total, all of which had typical circular structures consisting of a large single-copy region, a small single-copy region and a pair of inverted repeat regions. The comparative analysis showed that the chloroplast genome of P. paradoxum was conserved in the inverted repeat regions. Additionally, we identified 60 dispersed repeat sequences, 61-63 simple sequence repeats, and 30 codons within the 82 protein-coding genes that exhibited RSCU values greater than one. Furthermore, we detected highly divergent regions that hold potential as new DNA barcodes for species identification. Compared with Pseudogalium, the gene number, gene length, and GC content in the chloroplast genomes of Galium and Rubia exhibited differential characteristics, and the dispersed repeat sequences and SSRs in Galium and Rubia were significantly different. Phylogenetic analysis based on the whole chloroplast genomes showed that Pseudogalium can be treated as a new genus, with P. paradoxum subsp. paradoxum and P. paradoxum subsp. duthiei considered as two distinct subspecies of P. paradoxum.

Conclusions: The complete chloroplast genomes of P. paradoxum were first reported in this study, which provided a new insight into phylogeny and taxonomy of this genus. Phylogenetic analyses strongly supported the following proposals: (1) P. paradoxum can be isolated as a genus closely related to Galium; (2) P. paradoxum subsp. paradoxum and P. paradoxum subsp. duthiei form distinct clades, both of which can be considered as subspecies of P. paradoxum.

Keywords: Galium; Pseudogalium paradoxum; Rubia; Chloroplast genome; Comparative analysis; Phylogenetic relationships.

Fig. 1

Map of the chloroplast genome of Pseudogalium. Genes inside and outside the outer circle are transcribed in clockwise and counterclockwise directions, respectively. Genes belonging to different functional groups are colourcoded. The dark-gray region in the inner circle corresponds to GC content, whereas the lighter-gray region corresponds to AT content

Fig. 2

Types of dispersed repeat sequences in chloroplast genomes of Pseudogalium, Galium, and Rubia

Fig. 3

Simple sequence repeats (SSRs). A SSRs in Pseudogalium, Galium and Rubia. B Number of SSRs in LSC, IRs and SSC in Pseudogalium, Galium and Rubia. C Specific of SSRs in Pseudogalium. D Proportion of SSRs types in Pseudogalium

Fig. 4

Heatmap of all the RSCU values of amino acids and stop codons of 82 plastid protein-encoding genes. Red and blue values indicate higher and lower RSCU values, respectively

Fig. 5

Comparison of connection sites of LSC, IRb, SSC, and IRa in the chloroplast genomes. JLB (IRb/LSC), JSB (IRb/SSC), JSA (SSC/IRa), and JLA (IRb/ LSC) represent the junction sites between two adjacent regions in the genomes

Fig. 6

Sequence identity plots of the chloroplast genomes of Pseudogalium, Galium and Rubia

Fig. 7

Sliding window analyses of Pseudogalium, Galium and Rubia chloroplast genomes. The nucleotide diversity (Pi) value of each window is shown on Y-axis, and positions are shown on X-axis

Fig. 8

The complete chloroplast genomes comparison of Pseudogalium, Galium and Rubia with P. paradoxum subsp. paradoxum YangLE496 as a reference. The X-axis represents the coordinate in the chloroplast genome. The Y-axis shows different species names, and sequence similarity of aligned regions is displayed as horizontal bars, which expresses as a percentage within 50–100%

Fig. 9

Phylogenetic trees. (A) Phylogenetic tree of the Pseudogalium, Galium and Rubia inferred from ML analyses based on the complete chloroplast genomes. (B) Phylogenetic tree of the Pseudogalium, Galium and Rubia inferred from BI analyses based on 82 protein-coding genes in chloroplast genomes. (C) Phylogenetic tree of the Pseudogalium, Galium and Rubia based on the mutation hotspot regions in chloroplast genomes