# Comparative genomics and phylogenetic analysis of seven Ficus species based on chloroplast genomes

## TL;DR

This study compares chloroplast genomes of seven Ficus species to better understand their evolutionary relationships and identify useful genetic markers.

## Contribution

The study provides newly assembled and annotated chloroplast genomes for seven Ficus species, resolving gene content discrepancies and identifying hypervariable regions useful for DNA barcoding.

## Key findings

- The chloroplast genomes of seven Ficus species were assembled, showing a typical quadripartite structure and conserved gene content.
- Three hypervariable regions (ccsA, ccsA - ndhD, and rpoB - trnC-GCA) were identified as potential DNA barcodes for Ficus.
- Phylogenetic analysis using 79 protein-coding genes confirmed the monophyly of Ficus and grouped the seven species into two well-supported clades.

## Abstract

The genus Ficus (Moraceae) is a large and ecologically important group, known for its intricate fig-wasp pollination mutualism and role as a keystone resource in tropical ecosystems. Despite its significance, the phylogenetic relationships within Ficus remain partially unresolved, necessitating more comprehensive genomic data. Chloroplast (cp) genomes are valuable resources for plant phylogenetic and comparative genomic studies. Here, we sequenced, assembled, and comparatively analyzed the complete chloroplast genomes of seven Ficus species, including Ficus esquiroliana, Ficus pandurata, Ficus formosana, Ficus erecta, Ficus carica, Ficus hirta, and Ficus stenophylla.

The complete cp genomes were successfully assembled, ranging in size from 160,340 bp to 160,669 bp, and exhibited a typical quadripartite structure with highly conserved gene content and arrangement. Critically, while some of these species have previously published plastomes, our assemblies consistently encoded 130 genes, contrasting with reported gene counts (e.g., 129 for F. formosana (NC_059898), 119 for F. carica (KY635880), 131 for F. erecta (MT093220)) in earlier studies. Numerous repeat sequences and simple sequence repeats (SSRs) were identified, predominantly in non-coding regions, which serve as valuable resources for developing novel genetic markers. Analysis of codon usage revealed a strong bias towards A/T endings, a common feature in plant cp genomes. While inverted repeat (IR) boundary regions were largely conserved, minor variations, including partial gene duplications (rps19, rpl2), were observed. Comparative genome alignment and nucleotide diversity analysis showed high sequence conservation, with most variations concentrated in single-copy and non-coding regions. We identified three hypervariable regions (ccsA, ccsA - ndhD, and rpoB - trnC-GCA) with elevated nucleotide diversity (Pi > 0.012, ccsA up to 0.0141), suggesting their utility as candidate DNA barcodes for Ficus. Phylogenetic analysis using 79 protein-coding genes from 26 species robustly supported the monophyly of Ficus and resolved the seven newly sequenced species into two well-supported clades, consistent with previous classifications.

Our study provides new, consistently assembled and rigorously annotated chloroplast genome data for Ficus, including clarified data for previously studied species with notable gene content discrepancies. These data identify candidate molecular markers with potential applications for systematics and population genetics, and offer robust insights into relationships among sampled taxa. These data will facilitate future studies of Ficus evolution and conservation when complemented by broader taxon sampling and nuclear/mitochondrial data.