Phylogenetic Analysis and Codon Usage Bias of Chloroplast Genomes in Rhododendron

Objective The objective of this study was to analyze the codon usage patterns in the chloroplast genome of Rhododendron, aiming to provide genetic information for the conservation and innovation of Rhododendron germplasm resources and to lay a foundation for its genetic improvement and phylogenetic research.

Method The chloroplast genomes of 46 Rhododendron species were taken as the research objects. The phylogenetic relationships of Rhododendron were investigated using mafft and iQtree software. Software such as Codon W 1.4.2, R language, and EMBOSS were utilized to analyze parameters of codons in the chloroplast genome, including GC content, relative synonymous codon usage (RSCU), and effective number of codons (ENC). Additionally, neutrality plot, PR2-plot, and optimal codon analysis were carried out to explore the characteristics of codon usage preference and the main influencing factors in Rhododendron plants.

Result The phylogenetic tree of the 46 Rhododendron species revealed three major clades. Clade I comprised subgenera Hymenanthes and Pentanthera; Clade II mainly consisted of subgenus Rhododendron, including R. mucronulatum; and Clade III included subgenera Tsutsusi and Azaleastrum. The GC₁, GC₂, GC₃, and GC_all contents of Rhododendron plants were relatively close, and all species showed a consistent pattern of GC₁ > GC₂ > GC₃. The variation range of GC₃ content was smaller than that of GC₁ and GC₂, indicating the uneven distribution of GC content at different codon positions. The RSCU values of chloroplast genomes in Rhododendron ranged from 0.3123 to 2.156. The ENC values of chloroplast genomes in Rhododendron were mainly concentrated in the range of 35 to 55, indicating weak codon usage bias. The chloroplast genome codons of Rhododendron species tended to end with A or U. Codon usage bias was influenced by both mutation and natural selection, with natural selection playing a more important role. The codon usage bias of the chloroplast genome in subgenus Azaleastrum was more strongly influenced by natural selection than in other subgenera. A total of 7 optimal codons were identified in the chloroplast genomes of Rhododendron species, most of which end with A/U and primarily encode uncharged amino acids.

Conclusion The codon usage bias in the chloroplast genome of Rhododendron plants is mainly influenced by natural selection.