The MADS-box genes encode transcription factors with key roles in plant growth and development. A comprehensive analysis of the MADS-box gene family in bread wheat (Triticum aestivum) has not yet been conducted, and our understanding of their roles in stress is rather limited. Here, we report the identification and characterization of the MADS-box gene family in wheat. A total of 180 MADS-box genes classified as 32 Mα, 5 Mγ, 5 Mδ, and 138 MIKC types were identified. Evolutionary analysis of the orthologs among T. urartu, Aegilops tauschii and wheat as well as homeologous sequences analysis among the three sub-genomes in wheat revealed that gene loss and chromosomal rearrangements occurred during and/or after the origin of bread wheat. Forty wheat MADS-box genes that were expressed throughout the investigated tissues and development stages were identified. The genes that were regulated in response to both abiotic stresses (i.e., phosphorus deficiency, drought, heat, and combined drought and heat) and biotic stresses (i.e., Fusarium graminearum, Septoria tritici, stripe rust and powdery mildew) were detected as well. A few notable MADS-box genes were specifically expressed in a single tissue and those showed relatively higher expression differences between the stress and control treatment. The expression patterns of considerable MADS-box genes differed from those of their orthologs in Brachypodium, rice, and Arabidopsis. Collectively, the present study provides new insights into the possible roles of MADS-box genes in response to stresses and will be valuable for further functional studies of important candidate MADS-box genes.

Introduction: Plant factories provide a controlled platform for rice cultivation and rapid breeding, yet the effects of controlled environments on rice growth and yield formation remain poorly understood relative to field conditions. Methods: This study systematically compared growth duration, plant architecture, root and leaf traits, biomass accumulation, yield components, and environmental dynamics of three representative rice cultivars grown under field and plant factory conditions. Logistic model fitting was used to characterize plant height growth dynamics, and five machine learning models were further applied to predict plant height and assess the relative importance of growth- and environment-related variables. Results: The results showed that a plant factory shortened the average growth duration from 138 to 95 days compared with field cultivation and promoted early vegetative development, including stronger tillering, larger total leaf area, longer roots, and greater fresh biomass accumulation. It also increased the total panicles per unit area and the grain number per panicle, whereas seed-setting rate, 1000-grain weight, and final grain yield were not significantly increased. The machine learning models achieved high predictive accuracy for plant height (R ² >0.9), with growth duration as the dominant predictor in both systems, followed by CO ₂ concentration in the plant factory and cultivar in the field. Discussion: These findings reveal that controlled-environment cultivation reshapes rice developmental rhythm and vegetative-reproductive allocation, providing a physiological basis for understanding rice plasticity and optimizing plant factory-based rice production.

Background: Synonymous mutations have long been treated as neutral or silent variants and are therefore often filtered out or deprioritized in genetic analyses. Yet increasing evidence shows that, even without altering the protein sequence, they can influence gene function and phenotype through multiple RNA-level mechanisms. In plants, recent studies suggest that synonymous variants can act as causal alleles in domestication and agronomic traits. These developments suggest that synonymous variation represents an underappreciated component of plant causal genetics. Scope: This Perspective synthesizes current knowledge on functional synonymous variants in plants from a plant-centred perspective. We summarize the major mechanistic routes by which synonymous variants can become functional, curate representative plant cases spanning domestication, breeding and mutant discovery, and propose a Tier 0–3 evidence framework. We also outline a practical workflow for advancing candidate synonymous variants towards causal validation and discuss how these variants may be more explicitly incorporated into frameworks for causal inference and breeding deployment. Conclusions: Functional synonymous variants are not merely rare exceptions, but an emerging class of hidden causal alleles with implications for plant genetics, evolutionary biology and crop improvement. Reintegrating synonymous sites into the analytical framework of plant trait dissection should improve how genotype–phenotype relationships are interpreted and help reduce a persistent blind spot in candidate-variant prioritization. More broadly, synonymous variation may expand the set of causal alleles available for precision breeding by enabling fine phenotypic tuning without altering the protein sequence.

Herpetospermum pedunculosum is an endangered cucurbitaceous medicinal plant whose functional genomic and metabolic studies are constrained by the lack of efficient aseptic seedling and genetic transformation systems. Here, we established a rapid aseptic seedling production method and a RUBY-assisted hairy root transformation platform integrated with metabolomic evaluation. The cotton-supported liquid culture method provided a favorable balance between germination performance and experimental operability. Microsyringe-mediated stem-base inoculation with Rhizobium rhizogenes K599 carrying 35S::RUBY enabled rapid visual screening of RUBY-positive hairy roots, with a maximum transformation efficiency of 93.33%. Widely targeted metabolomics identified 242 metabolites and revealed major differences in amino acid/nitrogen and sugar metabolism, lipid-related metabolism, specialized metabolite biosynthesis, hormone-related pathways, and ABC transporters among wild-type, vector-free K599-induced, and RUBY-expressing roots. In addition, 80 HpRHL-like bHLH genes were identified. As a proof of concept, RT-qPCR confirmed HpRHL76 overexpression, which was associated with an increase in mean root hair length from approximately 0.4 to 1.1 mm. Together, this integrated framework provides a practical basis for preliminary candidate gene assessment, functional genomic studies, and future metabolic engineering of medicinally relevant compounds in H. pedunculosum, and may serve as a methodological reference for other difficult-to-transform medicinal plants.