Strigolactones (SLs) regulate plant shoot development by inhibiting axillary bud growth and branching. However, the role of SLs in wintersweet (Chimonanthus praecox) shoot branching remains unknown. Here, we identified and isolated two wintersweet genes, CCD7 and CCD8, involved in the SL biosynthetic pathway. Quantitative real-time PCR revealed that CpCCD7 and CpCCD8 were down-regulated in wintersweet during branching. When new shoots were formed, expression levels of CpCCD7 and CpCCD8 were almost the same as the control (un-decapitation). CpCCD7 was expressed in all tissues, with the highest expression in shoot tips and roots, while CpCCD8 showed the highest expression in roots. Both CpCCD7 and CpCCD8 localized to chloroplasts in Arabidopsis. CpCCD7 and CpCCD8 overexpression restored the phenotypes of branching mutant max3-9 and max4-1, respectively. CpCCD7 overexpression reduced the rosette branch number, whereas CpCCD8 overexpression lines showed no phenotypic differences compared with wild-type plants. Additionally, the expression of AtBRC1 was significantly up-regulated in transgenic lines, indicating that two CpCCD genes functioned similarly to the homologous genes of the Arabidopsis. Overall, our study demonstrates that CpCCD7 and CpCCD8 exhibit conserved functions in the CCD pathway, which controls shoot development in wintersweet. This research provides a molecular and theoretical basis for further understanding branch development in wintersweet.

Chimonanthus praecox, a member of the Calycanthaceae family, is a unique, traditional, and famous flowering economic tree species in China. Despite the existence of several varieties, only a few cultivars have been formally named. Currently, expression sequence tag–simple sequence repeat (EST-SSR) markers are extensively used to identify different species and varieties; a large number of microsatellites can be identified from transcriptome databases. A total of 162,638 unigenes were assembled using RNA-seq; 82,778 unigenes were annotated using the Nr, Nt, Swiss-Prot, Pfam, GO, KOG, and KEGG databases. In total, 13,556 SSR loci were detected from 11,691 unigenes, with trinucleotide repeat motifs being the most abundant among the six repeat motifs. To develop the markers, 64,440 pairs of SSR primers with polymorphism potential were designed, and 75 pairs of primers were randomly selected for amplification. Among these markers, seven pairs produced amplified fragments of the expected size with high polymorphism. Using these markers, 12 C. praecox varieties were clustered into two monophyletic clades. Microsatellites in the transcriptome of C. praecox exhibit rich types, strong specificity, and great polymorphism potential. These EST-SSR markers serve as molecular technical methods for identifying different varieties of C. praecox and facilitate the exploration of a large number of candidate genes associated with important traits.

To identify stable reference genes for qRT-PCR analysis across different developmental stages and color variations of tender leaves in Litsea coreana, seven candidate reference genes were selected based on existing transcriptome data. qRT-PCR was performed on tender leaves of L. coreana at various stages and under different color conditions. The stability of these genes was evaluated using GeNorm (version 2003), NormFinder (version 0953), BestKeeper (version 2003), and ReFinder software (version 2004). The most stable genes were selected, and the stability of the chosen reference genes was validated. RPL and ACT were the most stable genes across different leaf developmental stages, while ACT and EF1-α showed the highest stability across different leaf colors. Overall, ACT and EF1-α were the most stable reference genes for both developmental stages and color variations. ACT and EF1-α can be used as reliable reference genes for gene expression studies in the color change process of L. coreana tender leaves. This will provide a foundation for further research into the molecular mechanisms of leaf color changes and the development of color regulation genes in L. coreana.

TCP transcription factors represent a crucial family of plant regulators that contribute significantly to growth and developmental processes. Although the TCP gene family has been extensively studied in various plant species, research on Chimonanthus praecox (wintersweet) remains limited. Here, we performed genome-wide identification and analysis of the TCP gene family in C. praecox and identified 22 CpTCP genes. We further systematically examined the associated physicochemical properties, evolutionary relationships, gene structures, and regulatory features. Analysis revealed that all CpTCP proteins possess a conserved TCP domain, and subcellular localization prediction indicated their localization in the nucleus. Promoter analysis revealed that multiple cis-elements are associated with abiotic stress responses and plant growth regulation. Further analysis revealed high CpTCP2 expression in the leaves and stamen, with significantly increased levels during flower senescence. CpTCP2 expression was upregulated in response to methyl jasmonate (MeJA), salicylic acid, abscisic acid, and shade. CpTCP2 overexpression in Arabidopsis thaliana resulted in a reduced leaf area, delayed flowering, and increased rosette leaf numbers. Moreover, MeJA treatment accelerated leaf senescence in CpTCP2 transgenic Arabidopsis. These findings provide insights into the evolutionary characteristics of the TCP family in C. praecox, highlighting the functional role of CpTCP2 in regulating leaf development and flowering time in Arabidopsis, thereby offering valuable genetic resources for wintersweet molecular breeding.

Chimonanthus praecox, a traditional ornamental plant in China, is admired for its ability to bloom during the cold winter season and is recognized as an outstanding woody cut flower. MADS-box genes encode transcription factors essential for plant growth and development, with key functions in regulating flowering time and the formation of floral organs. In this study, 74 MADS-box genes (CpMADS1–CpMADS74) were identified and mapped across 11 chromosomes, with chromosome 1 harboring the highest number (13 genes) and chromosome 3 the fewest (3 genes). Physicochemical property analysis revealed that all CpMADS proteins are hydrophilic and predominantly nuclear-localized. Phylogenetic analysis classified these genes into Type I and Type II subfamilies, highlighting a clear divergence in domain structure. Eighty simple sequence repeat (SSR) loci were detected, with dinucleotide repeats being the most abundant, and the majority located in Type II MADS genes. From 23 C. praecox samples, 10 polymorphic SSR markers were successfully developed and PCR-validated, enabling a cluster analysis that grouped these cultivars into three distinct clusters. This study offers significant insights into the regulation of flowering, floral organ development, genetic linkage map construction, and the application of marker-assisted selection in C. praecox.