# Comparative chloroplast genome analysis of Polygala species: insights into evolution, phylogeny, and DNA barcoding

## TL;DR

This study analyzes the chloroplast genomes of Polygala species to understand their evolution, phylogeny, and potential for DNA barcoding.

## Contribution

The first report of the tetraploid Polygala vulgaris chloroplast genome and identification of hypervariable regions for DNA barcoding.

## Key findings

- The chloroplast genome sizes of Polygala species ranged from 165,246 bp to 168,779 bp with consistent gene content.
- Nine highly divergent genes and intergenic spacer regions were identified as potential DNA barcoding markers.
- Phylogenetic analysis revealed biogeographic disjunction and close relationship between P. vulgaris and P. amerella.

## Abstract

Polygala species are rich in saponin molecules and are traditionally used for medical purposes. Research on the chloroplast (cp) genome of Polygala is crucial to understanding its phylogenetics, biogeography, and species identification. In this study, the tetraploid Polygala vulgaris chloroplast genome was reported for the first time via various assembly and annotation tools. By combining these new data with the published cp genomes of 12 Polygala taxa from NCBI, we conducted a comparative analysis of nucleotide diversity, repeat sequences, genome synteny, phylogenetic relationships and DNA barcoding. The final assembled chloroplast genome sizes for the studied Polygala taxa ranged from 165,246 bp to 168,779 bp, with a total of 174 genes annotated, including 102 protein-coding genes, 63 transfer RNA genes, and 13 ribosomal RNA genes in Polygala vulgaris. A total of nine highly divergent genes were identified, including the CDS gene rpl32 as well as the intergenic spacer regions matK-trnK, trnL-trnF, trnH-psbA, trnQ-psbK, ndhI-ndhF, ndhF-rpl32, ccsA-ndhD, and ndhG-ndhI. Phylogenetic relationships were reconstructed based on Bayesian inference and maximum likelihood analyses using ortholog-based and concatenated datasets derived from the identified hypervariable regions separately. The topological structure of the constructed phylogenetic tree showed biogeographic disjunction, and the most similar relative of Polygala vulgaris was one of the European taxa, P. amerella, based on high support values. The analysis of relative synonymous codon usage revealed that a codon usage pattern largely consistent with that observed across other Polygala species. A total of three types of repeat sequences, forward, palindromic, and reverse, were identified across the chloroplast genomes of the analyzed species. These findings provide a valuable reference for phylogenomic and evolution of Polygala taxa as well as to develope molecular markers for DNA barcoding.