The complete chloroplast genome of Rhododendronambiguum and comparative genomics of related species.

Rhododendron Linnaeus, 1753, the largest genus of woody plants in the Northern Hemisphere, includes some of the most significant species in horticulture. Rhododendronambiguum Hemsl, 1911, a member of subsection Triflora Sleumer 1947, exemplifies typical alpine Rhododendron species. The analysis of the complete chloroplast genome of R.ambiguum offers new insights into the evolution of Rhododendron species and enhances the resolution of phylogenetic relationships. This genome is composed of 207,478 base pairs, including a pair of inverted repeats (IRs) of 47,249 bp each, separated by a large single-copy (LSC) region of 110,367 bp and a small single-copy (SSC) region of 2,613 bp. It contains 110 genes: 77 protein-coding genes, 29 tRNAs, four unique rRNAs (4.5S, 5S, 16S, and 23S), with 16 genes duplicated in the IRs. Comparative analyses reveal substantial diversity in the Rhododendron chloroplast genome structures, identifying a fourth variant pattern. Specifically, four highly divergent regions (trnI-rpoB, ndhE-psaC, rpl32-ndhF, rrn16S-trnI) were noted in the intergenic spacers. Additionally, 76 simple sequence repeats were identified. Positive selection signals were detected in four genes (cemA, rps4, rpl16, and rpl14), evidenced by high Ka/Ks ratios. Phylogenetic reconstruction based on two datasets (shared protein-coding genes and complete chloroplast genomes) suggests that R.ambiguum is closely related to R.concinnum Hemsley, 1889. However, the phylogenetic positions of subsection Triflora Pojarkova, 1952 species remain unresolved, indicating that the use of complete chloroplast genomes for phylogenetic research in Rhododendron requires careful consideration. Overall, our findings provide valuable genetic information that will enhance understanding of the evolution, molecular biology, and genetic improvement of Rhododendron spieces.