Chloroplasts (cp) are important organelles in plant cells that have been widely used in phylogenetic, molecular evolution, and gene expression studies due to their conserved molecular structure. In this study, we obtained the complete cp genome of Trivalvaria costata (Annonaceae) and analyzed its structural characteristics. Additionally, we analyzed the rps12 gene in the phylogenetic framework of magnoliids. The T. costata cp genome comprises 162,002 bp and contains 132 genes. We detected 48 simple sequence repeats (SSRs) and identified 29 high-frequency codons as well as 8 optimal codons. Our multiple analyses show that codon usage bias is mainly influenced by natural selection. For the first time, we found the rps12 gene to be entirely located in the IR region (in Annona). In groups with exon 1 located in the single-copy (SC) region and exons 2–3 located in the inverted repeat (IR) region, the transition rate and synonymous substitution rate of exon 1 were higher than those of exons 2–3. Adaptive evolution identified a positive selection site (116) located in the 3₁₀-helix region, suggesting that the rps12 gene may undergo adaptive changes during the evolutionary history of magnoliids. This study enhances our knowledge regarding genetic information on T. costata and provides support for reduced substitution rates in the IR region.

Plant chloroplast (cp) gene clusters consist of genes arranged closely together on the cp genome. These genes are organized in operon structures and participate in cotranscription, typically exhibiting conservation. Broken gene clusters have been observed in gymnosperms. In order to investigate whether the substitution rates and selection pressure of associated genes are affected following the disruption of gene clusters, the cp genomes of 80 species (78 gymnosperms and 2 outgroups) were analyzed. A phylogenetic analysis was conducted using 58 shared genes to examine the evolutionary rates and selection pressure of genes associated with gene clusters and protein-coding genes in Sciadopitys verticillata. The results demonstrate that S. verticillata exhibited the highest number of rearrangements compared to the Cycas revoluta genome. Four gene clusters (rps2, psbB, rpoB, and petL clusters) in S. verticillata were disrupted, while rps2 in Callitris rhomboidea experienced disruption. Significantly increased evolutionary rates were observed in 12 out of 18 gene cluster-related genes in S. verticillata. Following disruption, S. verticillata and C. rhomboidea exhibited an increase in gene cluster-related genes, particularly rps2, and higher selection pressure on both rps2 and atpA genes compared to other species. Furthermore, among the 58 genes shared by S. verticillata, the evolutionary rates of 36 genes increased, and the selection pressure on 13 genes exceeded that of other species. These results indicate an increased substitution rate of gene clusters in S. verticillata and C. rhomboidea. The large-scale rearrangement and elevated substitution rates of the cp genome in S. verticillata were revealed. This study sheds light on the heterogeneity of cp genome evolution in gymnosperms.

Anther is thought to release either bicellular or tricellular pollen when mature. Though a few species had been found to shed both bicellular and tricellular pollen, due to their low frequency, they had been overlooked as special cases of bicellular or tricellular pollen in previous phylogenetic studies. In the present work, the pollen cytologies of 89 species from 26 genera of Annonaceae were observed using the overall transparency method and paraffin sectioning method. The results show that 73 species from 25 genera distribute bicellular pollen, while 16 species from 10 genera shed both bicellular and tricellular pollen. This is the first time that so many species with both types of pollen has been observed in the same family. Combined with reports from other families, the plants that were known to shed both types of pollen included 15 families, 40 genera, and 52 species. Our results indicate that the coexistence of bicellular and tricellular pollen might be the third type of pollen cell number. And the systematic relationship among them is needed to be reanalyzed.