Rhododendron fortunei (Ericaceae) possesses valuable horticultural and medicinal values. However, the genomic information on R. fortunei is very limited. In this study, the complete chloroplast genome (cp) of R. fortunei was assembled and annotated, SSR loci were characterised, comparative genomic analysis was carried out, and phylogenetic research was also performed. The results showed that the R. fortunei cp genome was of a typical quadripartite structure (200,997 bp). The lengths of the large single copy region (LSC), the inverted repeat regions (IR), and the small single copy region (SSC) were 109,151 bp, 2,604 bp, and 44,619 bp, respectively. A total of 147 unique genes were identified, including 99 protein-coding genes, 42 tRNA genes, and 6 rRNA genes, respectively. Leucine (11.51%) and cysteine (1.15%) were the highest and lowest representative amino acids, respectively. The total of 30 codons with obvious codon usage bias were all A/U-ending codons. Among the 77 simple sequence repeats, the majority were mononucleotide A/T repeats located in the intergenic spacer region. Five gene regions showed high levels of nucleotide diversity (Pi > 0.03). The comparative genome analysis revealed 7 hotspot intergenic regions (trnI-rpoB, trnTrpl16, rpoA-psbJ, rps7-rrn16, ndhI-rps16, rps16-rps19, and rrn16-trnI), showing great potential as molecular makers for species authentication. Expansion and contraction were detected in the IR region of the R. fortunei cp genome. In the phylogenetic tree, R. fortunei was closely related to R. platypodum. This research will be beneficial for evolutionary and genetic diversity studies of R. fortunei and related species among the Ericaceae family.
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