Quantitative real-time PCR (qPCR) is an indispensable technique for gene expression analysis in modern molecular biology. The selection and evaluation of suitable reference genes is a prerequisite for accurate gene expression analysis. Schimasuperba is a valuable tree species that is environmentally adaptable and highly fire-resistant. In this study, 12 candidate reference genes were selected to check their stability of gene expression in different tissues under abiotic stresses: cold stress, salt stress, and drought stress by ΔCt, geNorm, NormFinder, BestKeeper, and RefFinder. The results indicated that AP-2 was the most stably expressed overall and for the cold stress and drought stress. eIF-5α gene expression was the most stable under the salt stress treatment, while UBQ expression was the most stable across mature leaves, shoots, stems, and roots. In contrast, UBC20, GAPDH, and TUB were the least stably expressed genes tested. This study delivers valid reference genes in S. superba under the different experimental conditions, providing an important resource for the subsequent elucidation of the abiotic stress adaptation mechanisms and genes with biological importance.

This work aimed to identify the chemical compounds of Cinnamomum burmannii leaf essential oil (CBLEO) and to unravel the antibacterial mechanism of CBLEO at the molecular level for developing antimicrobials. CBLEO had 37 volatile compounds with abundant borneol (28.40%) and showed good potential to control foodborne pathogens, of which Staphylococcus aureus had the greatest inhibition zone diameter (28.72 mm) with the lowest values of minimum inhibitory concentration (1.0 μg/mL) and bactericidal concentration (2.0 μg/mL). To unravel the antibacterial action of CBLEO on S. aureus, a dynamic exploration of antibacterial growth, material leakage, ROS formation, protein oxidation, cell morphology, and interaction with genome DNA was conducted on S. aureus exposed to CBLEO at different doses (1/2–2×MIC) and times (0–24 h), indicating that CBLEO acts as an inducer for ROS production and the oxidative stress of S. aureus. To highlight the antibacterial action of CBLEO on S. aureus at the molecular level, we performed a comparative association of ROS accumulation with some key virulence-related gene (sigB/agrA/sarA/icaA/cidA/rsbU) transcription, protease production, and biofilm formation in S. aureus subjected to CBLEO at different levels and times, revealing that CBLEO-induced oxidative stress caused transcript suppression of virulence regulators (RsbU and SigB) and its targeted genes, causing a protease level increase destined for the biofilm formation and growth inhibition of S. aureus, which may be a key bactericidal action. Our findings provide valuable information for studying the antibacterial mechanism of essential oil against pathogens.

In recent years, the field of biology has witnessed a surge of interest in genomics research due to the advancements in biotechnology. Gene expression pattern analysis plays a crucial role in this research, as it enables us to understand the regulatory mechanism of gene expression and the associated biological processes. Real-time quantitative polymerase chain reaction (q-PCR) is an efficient method to analyze the gene expression patterns, for which accuracy relies on the standardized analysis of reference genes. However, numerous studies have shown that no reference gene is universal in all conditions, so screening a suitable reference gene under certain conditions is of great importance. Cinnamomum burmannii (C. burmannii) is rich in volatile components and has high medicinal and economic value. However, knowledge of the screening of reference genes for the gene expression analysis of C. burmannii is insufficient. Aiming at this problem, we evaluated and screened the reference genes in C. burmannii under different experimental conditions, including different abiotic stresses (Cold-treated, PEG-treated and Nacl-treated), different tissues, leaves at different developmental stages and different chemical types. In this study, different algorithms (∆Ct, geNorm, NormFinder and BestKeeper) were used to evaluate the stability of the candidate reference genes, and RefFinder further merged the output data to screen out the optimum reference gene under various experimental conditions in C. burmannii. The results showed that the optimal reference gene number for gene standardization was 2 under different experimental conditions. RPL27|RPS15 was the most suitable combination under the Nacl-treated and PEG-treated samples. RPL27|APT was the optimum combination under the Cold-treated samples. The optimal combinations of other samples were EF1α|ACT7 for different tissues, eIF-5A|Gllα for different borneol clones in C. burmannii, RPS15|ACT7 for leaves at different developmental stages and RPS15|TATA for all samples. Additionally, two terpenoid synthesis-related genes (CbWRKY4 and CbDXS2) were standardized to verify the feasibility of the selected reference genes under different experimental conditions. This study will be helpful for the subsequent molecular genetic mechanism study of C. burmannii.

The intricate interplay between photosynthetic efficiency and terpenoid biosynthesis in plants remains a pivotal yet underexplored area in secondary metabolism research. This study elucidates the physiological and molecular mechanisms underlying this synergy in Cinnamomum burmanni, a chemically diverse Lauraceae species, through a multi-omics approach. A high-quality chromosome-level genome of C. burmanni (1.14 Gb, scaffold N50: 94.90 Mb) was assembled, and evolutionary analyses revealed species-specific gene family expansions, particularly in mono-, sesqui-, and diterpenoid biosynthesis pathways. Comparative analyses of photosynthetic traits across chemotypes demonstrated that the borneol-type exhibits superior photosynthetic capacity, characterized by elevated net photosynthetic rate, transpiration rate, stomatal conductance, intercellular CO₂, carboxylation efficiency and non-photochemical quenching. The phenotype is linked to upregulated chlorophyll metabolism, carotenoid biosynthesis regulators, and enhanced light-harvesting complex and photosystem components, optimizing light energy conversion. Mechanistically, photosynthetic activity modulates precursor flux into terpenoid pathways by regulating rate-limiting enzymes. Additionally, lineage-specific expansions of terpene synthase and isopentenyl diphosphate synthase gene families underpin specialized terpenoid production. A post-transcriptional regulatory network involving 14 miRNAs (e.g., miR396, miR2950) was identified, coordinately targeting 11 key genes in both photosynthesis and terpenoid synthesis, suggesting a dual role in metabolic fine-tuning. This work advances understanding of the evolutionary and physiological integration of photosynthesis and secondary metabolism in aromatic plants, offering a genomic foundation for biotechnological applications in metabolite synthesis and chemotype breeding.

Schima superba, commonly known as the Chinese guger tree, is highly adaptable and tolerant of poor soil conditions. It is one of the primary species forming the evergreen broad-leaved forests in southern China. Dirigent proteins (DIRs) play crucial roles in the synthesis of plant lignin and lignans, secondary metabolism, and response to adversity stress. However, research on the DIR gene family in S. superba is currently limited. This study identified 24 SsDIR genes, categorizing them into three subfamilies. These genes are unevenly distributed across 13 chromosomes, with 83% being intronless. Collinearity analysis indicated that tandem duplication played a more significant role in the expansion of the gene family compared to segmental duplication. Additionally, we analyzed the expression patterns of SsDIRs in different tissues of S. superba. The SsDIR genes exhibited distinct expression patterns across various tissues, with most being specifically expressed in the roots. Further screening identified SsDIR genes that may regulate drought stress, with many showing differential expression under drought stress conditions. In the promoter regions of SsDIRs, various cis-regulatory elements involved in developmental regulation, hormone response, and stress response were identified, which may be closely related to their diverse regulatory functions. This study will contribute to the further functional identification of SsDIR genes, providing insights into the biosynthetic pathways of lignin and lignans and the mechanisms of plant stress resistance.

The essential oil of Cinnamomum burmannii (Nees and T. Nees) Blume is rich in monoterpenes and sesquiterpenes. The post-transcriptional regulatory mechanisms controlling the expression of terpenoid-related genes have not yet been clarified in C. burmannii. Here, we conducted a metabolomic analysis of the leaves of C. burmannii across four developmental stages using gas chromatography–mass spectrometry. We also identified miRNAs and their target genes involved in terpenoid biosynthesis using small RNA sequencing. A total of 135 differentially expressed metabolites were detected, including 65 terpenoids, 15 aldehydes, and 13 benzenes. A total of 876 miRNAs from 148 families were detected, among which 434 miRNAs were differentially expressed, including three known miRNAs and 431 novel miRNAs. Four miRNAs (gma-miR5368, novel_miR_377, novel_miR_111, and novel_miR_251) were predicted to regulate the expression of four differential expressed genes involved in the monoterpenoid and sesquiterpenoid synthesis. miRNAs families miR396, miR5185, and miR9408 were predicted to play diverse regulatory roles in monoterpenoid and sesquiterpenoid synthesis during the leaf development of C. burmannii. The results of our study shed new light on the roles of regulatory genes in terpenoid biosynthesis. Our findings also have implications for the further promotion of essential oil production using the leaves of C. burmannii.