Self-incompatibility (SI) is an important plant mechanism that prevents inbreeding depression by recognizing and rejecting self-pollen, thereby promoting outcrossing. However, SI can also act as a barrier in breeding programs, presenting significant challenges to breeders. Passion fruit (Passiflora edulis), a tropical fruit species of substantial economic importance, also serves as a valuable system for investigating SI mechanisms within the Passifloraceae. Nevertheless, the molecular basis of SI in passion fruit has not yet been elucidated. In this study, we investigated the SI system in yellow passion fruit (P. edulis f. flavicarpa) and employed transcriptomic analysis to examine the time-course transcriptional responses following different pollination treatments. Transcriptomic analysis revealed distinct gene expression dynamics under different pollination treatments: self-pollinated samples exhibited stronger and earlier transcriptional changes, whereas the number of differentially expressed genes (DEGs) in cross-pollinated samples was relatively lower. Numerous pathways previously associated with sporophytic self-incompatibility (SSI) were enriched in the stigma samples after self-pollination. Reactive oxygen species (ROS) are crucial signaling molecules involved in pollen germination and pollen tube growth during SI responses. Our results showed that ROS-related pathways were enriched in stigma tissues after self-pollination. In addition, oxidative stress-related responses were detected in the style shortly after self-pollination, suggesting that plastid-associated or general oxidative stress processes may also be involved, although the precise source of ROS requires further validation. FERONIA, ROP9, and ARC1 are key genes related to the SI system in Brassica. In the passion fruit SI response, the expression levels of these genes increased in the style, indicating a spatial expression pattern different from that reported in classical Brassicaceae SSI systems. Together with cytological observations showing that self-pollen rejection occurs at the stigma surface, our results suggest that yellow passion fruit may employ an SSI-like regulatory framework while exhibiting a lineage-specific spatial deployment of SI-related regulators. Overall, this study provides new transcriptomic insights into the SI mechanism of yellow passion fruit, establishes a molecular framework for understanding SI in P. edulis f. flavicarpa, and offers novel insights into the diversity of plant SI systems.

Cyclic nucleotide-gated channel (CNGC) genes play key regulatory roles in plant immunity and abiotic stress responses. In this study, we conducted a genome-wide identification and analysis of the CNGC gene family in Suaeda glauca. A total of 44 SgCNGC genes were identified. Through phylogenetic analysis, gene structure analysis, chromosome distribution, conserved motif analysis, collinearity analysis, cis-acting element analysis, subcellular localization, and gene overexpression analysis, we systematically characterized the evolutionary relationships, structural features, and potential functions of this gene family. The results indicate that the SgCNGC gene family is evolutionarily highly conserved but exhibits functional divergence in structure and expression. Furthermore, functional assays revealed that overexpression of SgCNGC13 in Arabidopsis thaliana led to increased salt sensitivity, indicating a negative regulatory role for this gene under salt stress. These findings provide a foundation for understanding the role of the CNGC gene family in the growth, development, and stress response of S. glauca and contribute to the remediation of saline-alkali land.

Auxin plays a central role in coordinating floral organ differentiation and fruit development, yet how auxin signaling is coupled with auxin metabolic homeostasis during pineapple reproductive development remains poorly understood. Here, we systematically characterized the GH3 gene family in pineapple (Ananas comosus) and integrated these data with 20 AcoARF members to investigate the organization and function of ARF–GH3 regulatory modules. Seventeen AcoGH3 genes were identified, and phylogenetic and collinearity analyses revealed both evolutionary conservation and diversification of the ARF and GH3 families. Promoter analysis further showed that auxin response elements (AuxREs) were widely but differentially distributed among AcoGH3 promoters, providing a cis-regulatory basis for selective ARF recognition. Transcriptomic profiling across floral organs, ovaries, gynoecia, and developing fruits revealed pronounced tissue- and stage-specific expression patterns, indicating dynamic reorganization of ARF-GH3 modules during reproductive development. Yeast one-hybrid assays demonstrated selective binding of AcoARFs to AuxRE-containing AcoGH3 promoters, while dual-luciferase assays revealed distinct transcriptional outputs. Notably, AcoARF8 directly recognized and significantly activated the promoters of AcoGH3.1 and AcoGH3.3, whereas AcoARF4 repressed and AcoARF15 activated AcoGH3.14. Integration of developmental expression patterns with these molecular assays identified AcoARF8 as a prominent positive regulator of GH3 transcription during petal development. We therefore propose a developmental model in which AcoARF8 activity progressively increases during petal maturation, leading to enhanced transcription of AcoGH3.1 and AcoGH3.3 through AuxRE-dependent promoter regulation. Together, these findings establish a dynamic ARF–GH3 regulatory framework linking auxin-responsive transcription with auxin homeostasis and provide new insights into the molecular regulation of floral organ and fruit development in pineapple.

UDP-dependent glycosyltransferases (UGTs) are a large and diverse enzyme family involved in the modification and diversification of plant secondary metabolites, including flavonoids associated with fruit nutritional value and quality traits. However, the evolutionary organization of UGT families and their associations with flavonoid metabolism remain insufficiently characterized in tropical monocot fruit crops. Here, we performed a genome-wide characterization of the UGT family in pineapple (Ananas comosus) and integrated transcriptomic and metabolomic information to prioritize AcUGT candidates associated with flavonoid accumulation. A total of 68 AcUGT genes were identified and classified into 16 phylogenetic clades. Comparative genomic analyses demonstrated conserved syntenic relationships with other monocots, while tandem and segmental duplication contributed to AcUGT family expansion under predominant purifying selection. Structural analyses revealed conservation of the C-terminal plant secondary product glycosyltransferase (PSPG) motif involved in UDP-sugar recognition, whereas N-terminal sequence diversification distinguished AcUGT members. Promoter analysis identified cis-regulatory elements associated with hormone, developmental, environmental, and secondary metabolism responses. Integration of transcriptomic and metabolomic datasets revealed stage- and cultivar-associated AcUGT expression patterns that were statistically associated with flavonoid profiles. Candidate prioritization based on phylogeny, developmental or cultivar specificity and transcript–metabolite associations highlighted AcUGT29, AcUGT02, and AcUGT51 as high-priority candidates for future functional investigation. This study provides a genomic and metabolic framework for investigating UGT-associated flavonoid diversification and for guiding subsequent functional and fruit-quality studies in pineapple.

Ipomoea pes-caprae (IPC) is a perennial halophytic vine with remarkable salt and drought tolerance, playing a critical ecological and medicinal role in tropical and subtropical coastal ecosystems. Despite the availability of a high-quality chromosome-level reference genome and abundant transcriptome data, the absence of an integrated data platform has hindered in-depth functional gene discovery and genomic research in IPC. To address this gap, we developed the IPC Genome Database (IGD), the first comprehensive multi-omics database dedicated to IPC. IGD provides high-quality genomic sequences, gene structure annotations, and functional annotations, along with transcriptomic expression profiles under salt stress across different tissues. It also includes time-course expression data of roots and leaves under salt stress treatment, as well as leaf expression profiles under cold and heat stress conditions. The platform also integrates a suite of user-friendly, interactive bioinformatics analysis and visualization tools to facilitate efficient data exploration and functional mining. In particular, we established a specialized subset of stress-responsive genes to support rapid candidate gene screening and evolutionary analysis related to IPC’s salt and stress tolerance traits. By implementing automated data integration and quality control pipelines tailored for non-model species, IGD overcomes technical challenges such as fragmented data and delayed updates. As a centralized and scalable resource, IGD significantly lowers the barrier to functional genomics research in IPC and other halophytes, promoting their application in genetic improvement and ecological sustainability. In the future, IGD will continue to incorporate more omics data from halophytic species, further enhancing its value in salt tolerance research and molecular breeding efforts.

Caffeic acid O-methyltransferase (COMT) catalyzes the penultimate methylation in monolignol biosynthesis, controlling lignin composition and abiotic-stress tolerance. Kenaf (Hibiscus cannabinus L.), a fast bast-fiber crop rich in lignin, is valued for its mechanical strength and resilience to salinity. However, the COMT gene family has not yet been systematically characterized in this species. Here, we integrated phylogenetics, synteny, promoter and transcriptome analyses to create a comprehensive profile of kenaf COMT genes. Genome-wide screening identified 81 HcCOMT genes. Phylogenetic reconstruction with COMTs from Arabidopsis thaliana and Gossypium hirsutum resolved 10 distinct clades. Synteny analysis revealed 2 collinear blocks with Arabidopsis and 14 with cotton, whereas intraspecific duplication events indicated recent lineage-specific expansion. Promoter analysis identified numerous cis-elements responsive to light, phytohormones and abiotic stress, suggesting complex transcriptional regulation. Transcriptome mining uncovered 6 candidate genes with pronounced tissue specificity and salt responsiveness; qRT-PCR confirmed these patterns in root, stem and leaf tissues under 200 mM NaCl: HcCOMT28 and HcCOMT29 were repressed in the leaf, whereas HcCOMT11, HcCOMT12, HcCOMT13, and HcCOMT17 were up-regulated, consistent with altered lignin deposition patterns. Our findings provide a comprehensive genomic resource delineating the structure, evolution, and salt-responsive expression of the kenaf COMT family, and establish a foundation for elucidating the molecular mechanisms underlying lignin-mediated salt tolerance and for breeding elite kenaf cultivars with tailored fiber properties.

Nitrogen (N) is crucial for plant growth and stress resistance and is primarily absorbed and transported by nitrate transporters (NRT). Suaeda glauca, known for its strong salt-alkali stress resistance, and SgNRT genes have rarely been reported. This study aims to identify and analyze the SgNRT gene family to understand its composition, evolutionary patterns, and roles in salt stress responses. We identified 212 SgNRTs, which were categorized into three branches, with SgNRT1/SgNPF and SgNRT2 as the major families. Structural analysis, conserved domains, chromosomal localization, and collinearity were also examined. Spatiotemporal expression characteristics of SgNRT genes were analyzed, revealing specific expression across 13 organs or tissues and dynamic responses to salt treatment over 48 h. Notably, SgNRT1.185, SgNRT2.25, and SgNRT2.2 exhibited rapid salt induction in leaves (activated within 0.5 h, peaking at 2 h), with SgNRT1.185 showing relatively high upregulation. SgNRT1.185 and SgNRT2.35 were induced by high salt concentrations (200 mM) in both roots and leaves. SgNRT2.35 exhibited higher basal and stress-induced levels than the other genes. Bioinformatics analysis suggests spatially specific expression of SgNRT genes, potentially involved in nitrogen absorption and transport across various developmental stages and organs/tissues of Suaeda glauca. These findings offer a theoretical basis for understanding the adaptive strategies of Suaeda glauca under saline-alkali stress and provide insights into the functional evolution of plant NRT genes, aiding in the development of stress-resistant crops.

The cell cycle is a fundamental process of plant growth, development, and reproduction, in which cyclin-dependent kinases (CDKs) and cyclins (CYCs) play central roles in regulating the progression through various stages. These proteins are coordinated with multiple interacting partners to ensure the accurate execution of essential biological events such as DNA replication, chromosome segregation, and cell division. Marchantia polymorpha, one of the earliest diverging land plant species, has emerged as a key model for exploring fundamental mechanisms in plant biology and evolution. However, compared with other model plants, such as Arabidopsis thaliana and Oryza sativa, the core cell cycle genes in M. polymorpha remain relatively uncharacterized. In this study, we identified 31 core cell cycle genes in M. polymorpha through genome-wide analysis, including 13 CDKs, 8 CYCs, 5 E2F/DPs, 1 ICK, 1 RB, 1 CKS, and 2 Wee1 genes. We further analyzed their physicochemical properties, gene structures, and conserved domains, along with evolutionary pressures assessed via Ka/Ks and 4DTv analyses. Comparative genomic analysis revealed patterns of gene contraction and expansion. Additionally, we predicted cis-acting regulatory elements and performed differential expression analysis under various stress conditions to explore their potential functions and expression profiles. Finally, a protein-protein interaction (PPI) network was constructed, and key genes were experimentally validated. These findings provide valuable insights into the core cell cycle gene family in M. polymorpha, contributing to an enhanced understanding of cell cycle regulation and its evolutionary significance in plants.

TALE (Three Amino acid Loop Extension) homeodomain transcription factors are key conserved elements in eukaryotic developmental patterning. In plants, this superclass divides into the KNOX and BELL families, which are essential for regulating meristem maintenance, organogenesis, and tissue identity. Recent advances show that TALE proteins are intricately involved in plant reproductive processes, including gametophyte differentiation, embryonic axis formation, and floral organogenesis. They function as molecular scaffolds, integrating spatiotemporal signals and hormonal signaling like auxin, cytokinin, and gibberellin to control phase transitions and reproductive cell fate determination. The lineage-specific expansions and domain rearrangements of TALE genes across bryophytes, gymnosperms, and angiosperms indicate repeated co-option and neofunctionalization throughout land plant evolution. Emerging insights from epigenomics and protein interactomes reveal that TALE complexes modulate cell type-specific transcriptional responses. This review synthesizes current understanding of TALE-mediated regulatory networks during plant reproductive development and presents a conceptual framework for investigating their roles in developmental plasticity and stress-responsive fertility. We also highlight opportunities to utilize TALE-based regulatory modules to develop climate-resilient crops through multi-omics and genome editing approaches. Decoding the reproductive logic embedded in TALE networks offers transformative potential for reprogramming plant development in an era of agricultural and ecological uncertainty.

Passion fruit (Passiflora edulis), a tropical crop of significant economic value, exhibits temperature-sensitive floral development. Here, we identified 23 TALE transcription factors (PeTALEs) and characterized their roles in floral organogenesis and thermal adaptation. Phylogenetic analysis classified PeTALEs into KNOX and BELL subfamilies, with conserved domain architectures and cis-regulatory elements linked to stress and hormone signaling. Spatiotemporal expression profiling revealed PeTALE21 as a key regulator of corona initiation, while PeTALE17 dominated in later floral stages. Temperature stress assays demonstrated cold-induced upregulation of PeTALE15/16/19/22 and heat-mediated suppression of PeTALE10/18/21. Yeast two-hybrid assays uncovered functional interactions between PeTALE3/16/18/22/23, highlighting a network governing floral thermoresilience. This study provides the first genome-wide analysis of PeTALEs, offering insights for breeding climate-resilient passion fruit varieties.

Stress-associated proteins (SAPs), characterized by zinc finger domains, play a crucial role in regulating plant responses to various stresses. These proteins modulate stress-related gene expression and are integral to enhancing plant immunity, development, cell proliferation, and hormone regulation. In this study, we conducted a genome-wide analysis of the SAP gene family in Suaeda glauca (S. glauca), identifying 15 SAP genes encoding A20/AN1 zinc finger proteins. Functional analyses of three candidate genes under salinity stress were performed, examining phenotypic and physiological responses to better understand their role in stress tolerance. Sequence alignment, conserved domain analysis, and gene structure analysis revealed high conservation among S. glauca SAPs. Phylogenetic analysis identified two major groups within the gene family, providing insights into their evolutionary relationships. Transcription profiling analysis demonstrated significant expression of most SAP genes in response to salt stress, with qPCR validation confirming the upregulation of specific genes. Notably, transgenic Arabidopsis lines heterologously overexpressing the candidate genes SgSAP4, SgSAP5, and SgSAP7 demonstrated enhanced tolerance to salinity stress. This was evident from improved seed germination, root elongation, and reduced levels of stress markers, including malondialdehyde and free proline, compared to wild-type plants. These findings highlight the potential of these SAP genes in breeding programs aimed at improving salinity tolerance in crops.

Anthracnose of the tea plant (Camellia sinensis), caused by Colletotrichum spp., poses a significant threat to both the yield and quality of tea production. To address this challenge, researchers have looked to the application of endophytic bacteria as a natural alternative to the use chemical pesticides, offering potential for enhancing disease resistance and abiotic stress tolerance in tea plants. This study focused on identifying effective microbial agents to combat tea anthracnose caused by Colletotrichum fructicola. A total of 38 Bacillus-like strains were isolated from the tea rhizosphere, with 8 isolates showing substantial inhibitory effects against the mycelial growth of C. fructicola, achieving an average inhibition rate of 60.68%. Among these, strain T3 was particularly effective, with a 69.86% inhibition rate. Through morphological, physiological, and biochemical characterization, along with 16S rRNA gene phylogenetics analysis, these strains were identified as B. inaquosorum (T1 and T2), B. tequilensis (T3, T5, T7, T8, and T19), and B. spizizenii (T6). Biological and molecular assays confirmed that these strains could induce the expression of genes associated with antimicrobial compounds like iturin, fengycin, subtilosin, and alkaline protease, which effectively reduced the disease index of tea anthracnose and enhanced tea plant growth. In conclusion, this study demonstrates that B. inaquosorum, B. tequilensis, and B. spizizenii strains are promising biocontrol agents for managing tea anthracnose.

Heat stress transcription factors (HSFs) are the major regulators of plant response to environmental stress, especially heat and drought stress. To gain a deeper understanding of the mechanisms underlying HSFs in the abiotic stress response of passion fruit, we conducted an in silico analysis of the HSF gene family. Through bioinformatics and phylogenetic analyses, we identified 18 PeHSF members and classified them into A, B, and C groups. Collinearity analysis results revealed that the expansion of the PeHSF gene family was due to the presence of segmental duplication. Furthermore, gene structure and protein domain analysis illustrated that PeHSFs in the same subgroup are relatively conserved. Conserved motif and function domain analysis suggested that PeHSF proteins possess typical conserved functional domains of the HSF family. A protein interaction network and 3D structure prediction were used to study the potential regulatory relationship of PeHSFs. Additionally, the subcellular localization results of PeHSF-A6a, PeHSF-B4b, and PeHSF-C1a were consistent with the predictions. RNA-seq and RT-qPCR analysis revealed the expression patterns of PeHSFs in different tissues of passion fruit floral organs. Promoter analysis and the expression patterns of the PeHSFs under different treatments demonstrated their involvement in various abiotic stress processes. Notably, overexpression of PeHSF-C1a consistently enhanced tolerance to drought and heat stress in Arabidopsis. Overall, our findings provide a scientific basis for further functional studies of PeHSFs that could contribute to improvement of passion fruit breeding.

In plants, sexual reproduction relies on the proper development of floral organs that facilitate the successful development of fruits and seeds. Auxin responsive small auxin-up RNA (SAUR) genes play essential roles in floral organ formation and fruit development. However, little is known about the role of SAUR genes in pineapple floral organ formation and fruit development as well as stress responses. In this study, based on genome information and transcriptome datasets, 52 AcoSAUR genes were identified and grouped into 12 groups. The gene structure analysis revealed that most AcoSAUR genes did not have introns, although auxin-acting elements were abundant in the promoter region of AcoSAUR members. The expression analysis across the multiple flower and fruit development stages revealed differential expression of AcoSAUR genes, indicating a tissue and stage-specific function of AcoSAURs. Correlation analysis and pairwise comparisons between gene expression and tissue specificity identified stamen-, petal-, ovule-, and fruit-specific AcoSAURs involved in pineapple floral organs (AcoSAUR4/5/15/17/19) and fruit development (AcoSAUR6/11/36/50). RT-qPCR analysis revealed that AcoSAUR12/24/50 played positive roles in response to the salinity and drought treatment. This work provides an abundant genomic resource for functional analysis of AcoSAUR genes during the pineapple floral organs and fruit development stages. It also highlights the role of auxin signaling involved in pineapple reproductive organ growth.

In plants, sugars are required for several essential functions, including growth, storage, signaling, defense and reproduction. Sugar transporters carry out the controlled movement of sugars from source (leaves) to sink (fruits and roots) tissues and determine the overall development of the plant. Various types of sugar transporter families have been described in plants, including sucrose transporters (SUC/SUT), monosaccharide transporter (MST) and SWEET (from "Sugar Will Eventually be Exported Transporters"). However, the information about pineapple sugar transporters is minimal. This study systematically identified and classified 45 MST and 4 SUC/SUT genes in the pineapple genome. We found that the expression patterns of sugar transporter genes have a spatiotemporal expression in reproductive and vegetative tissues indicating their pivotal role in reproductive growth and development. Besides, different families of sugar transporters have a diel expression pattern in photosynthetic and non-photosynthetic tissues displaying circadian rhythm associated participation of sugar transporters in the CAM pathway. Moreover, regulation of the stress-related sugar transporters during cold stress indicates their contribution to cold tolerance in pineapple. Heterologous expression (yeast complementation assays) of sugar transporters in a mutant yeast strain suggested that SUT1/2 have the ability to transport sucrose, and STP13, STP26, pGlcT-L2 and TMT4 are able to transport glucose, whereas SWEET11/13 transport both sucrose and fructose. The information provided here would help researchers further explore the underlying molecular mechanism involved in the sugar metabolism of pineapple.

The WRKY transcription factors (TFs) are among the most diverse TF families of plants. They are implicated in various processes related to plant growth and stress response. Kenaf (Hibiscus cannabinus L.), an important fiber crop, has many applications, including the phytoremediation of saline-alkaline soil. However, the roles of WRKY TFs in kenaf are rarely studied. In the present study, 46 kenaf WRKY genes were genome-widely identified and characterized by gene structure, phylogeny and expression pattern analysis. Furthermore, the HcWRKY44 gene was functionally characterized in Arabidopsis under salinity and drought stresses. HcWRKY44 is a nuclear-localized protein that is positively induced by salinity and drought, with roots showing maximum accumulation of its transcripts. Under NaCl and abscisic acid (ABA) stress conditions, plants overexpressing HcWRKY44 had higher germination rates, better root growth and increased survival than control plants; however, it did not improve the ability to withstand drought stress. Moreover, ABA signaling genes (ABI1, ABI2, and ABI5), ABA-responsive genes (ABF4, RD29B, COR15A, COR47, and RD22), stress-related genes (STZ, P5CS, and KIN1), and ionic homeostasis-related genes (SOS1, AHA1, AHA2, and HKT1) were positively induced in HcWRKY44 transgenic plants under NaCl treatment. These results suggest that HcWRKY44 improved plant’s tolerance to salt stress but not osmotic stress through an ABA-mediated pathway. In summary, this study provides provided comprehensive information about HcWRKY genes and revealed that HcWRKY44 is involved in salinity tolerance and ABA signaling.

Kenaf (Hibiscus cannabinus L.) is an environmentally friendly, multipurpose fiber crop suitable for osmotic stress tolerance studies. However, the mechanisms of tolerance remain largely unknown. Here, we identified a stress-responsive HcWRKY50 gene from kenaf (Hibiscus cannabinus L.) and studied its function and tolerance under drought stress. HcWRKY50 is a nuclear-localized protein. The overexpression of HcWRKY50 in Arabidopsis showed higher drought tolerance, exhibiting increased root length and lateral root number, and reduced stomatal density compared with the control lines. The seed germination and seedling growth of HcWRKY50 transgenic plants showed less sensitivity to ABA but they became more sensitive to ABA in their stomatal aperture. Furthermore, qRT-PCR analysis revealed that HcWRKY50 regulated ABA signaling by promoting the expression of several key ABA-responsive and stress-responsive genes such as RD29B and COR47 in transgenic lines. Taken together, this study demonstrated that the kenaf transcription factor HcWRKY50 regulates seed germination and seedling growth and improves drought stress tolerance via an ABA signaling pathway.

BACKGROUND:Cell polarity establishment and maintenance is indispensable for plant growth and development. In plants, the YABBY transcription factor family has a distinct role in leaf asymmetric polarity establishment and lateral organ initiation. However, for the important sugar crop Saccharum, little information on YABBY genes is available.RESULTS:In this study, a total of 20 sequences for 7 SsYABBY genes were identified in the sugarcane genome, designated as SsYABBY1-7 based on their chromosome locations, and characterized by phylogenetic analysis. We provided a high-resolution map of SsYABBYs' global expression dynamics during vegetative and reproductive organ morphogenesis and revealed that SsYABBY3/4/5 are predominately expressed at the seedling stage of stem and leaf basal zone; SsYABBY2/5/7 are highly expressed in ovules. Besides, cross-species overexpression and/or complementation verified the conserved function of SsYABBY2 in establishing leaf adaxial-abaxial polarity and ovules development. We found that the SsYABBY2 could successfully rescue the leaves curling, carpel dehiscence, and ovule abortion defects in Arabidopsis crc mutant.CONCLUSIONS:Collectively, our study demonstrates that SsYABBY genes retained a conserved function in establishing and preserving leaf adaxial-abaxial polarity and lateral organ development during evolution.

Background Sugarcane (Saccharum spontaneum L.), the major sugar and biofuel feedstock crop, is cultivated mainly by vegetative propagation worldwide due to the infertility of female reproductive organs resulting in the reduction of quality and output of sugar. Deciphering the gene expression profile during ovule development will improve our understanding of the complications underlying sexual reproduction in sugarcane. Optimal reference genes are essential for elucidating the expression pattern of a given gene by quantitative real-time PCR (qRT-PCR). Method In this study, based on transcriptome data obtained from sugarcane ovule, eighteen candidate reference genes were identified, cloned, and their expression levels were evaluated across five developmental stages ovule (AC, MMC, Meiosis, Mitosis, and Mature). Results Our results indicated that FAB2 and MOR1 were the most stably expressed genes during sugarcane female gametophyte development. Moreover, two genes, cell cycle-related genes REC8 and CDK, were selected, and their feasibility was validated. This study provides important insights into the female gametophyte development of sugarcane and reports novel reference genes for gene expression research on sugarcane sexual reproduction.