<b>Chloroplast genome evolution of </b><b><i>Berberis </i></b><b>(Berberidaceae): implications for phylogeny and metabarcoding</b>

<i>Berberis</i> species play a crucial role in the formulation of Chinese patent medicine, Berberidis Radix, employed to address a range of ailments. These species have similar morphology, but different chemicals, potentially leading to misidentifications that can impact the medicine's efficacy. Therefore, it is imperative to develop suitable molecular markers to identify medicinal species. Furthermore, discrepancies exist in the taxonomy of the <i>Berberis</i> genus. In the current study, we <i>de novo</i> assembled the chloroplast genome of those six <i>Berberis</i> species (<i>B. </i><i>julianae</i>, <i>B. tsienii</i>, <i>B. pruinose</i>, <i>B. thunbergii</i>, <i>B. poiretii</i>, and <i>B. wilsoniae</i>) that commonly constitute Berberidis Radix, and compared them with other reported genomes. Our comparative analysis revealed similarities in genome structure, relative synonymous codon usage, amino acid frequency, repeats, and substitutions. Inverted repeats expansion and contraction affected the rate of mutations and found link to the phylogenetic classification of <i>Berberis</i>. We identified the <i>ndhD-ccsA</i> region as the most polymorphic region and applied it to the metabarcoding of Berberidis Radix medicine. The metabarcoding provided insight into their biological composition and confirmed six <i>Berberis </i>species commonly constitute Berberidis Radix. However, we also identified other <i>Berberis </i>species with lower read abundance. Our phylogenetic analysis supports the division of <i>Berberis</i> into four genera, which corroborates previous studies involving extensive sampling. In conclusion, this study offers comprehensive insights into the molecular evolution of the chloroplast genome and the phylogeny of the <i>Berberis</i> genus. Concurrently, through metabarcoding, we confirm the biological origin of Berberidis Radix, thus shedding light on its composition.