Dendrobine, a neuroprotective and anticancer sesquiterpenic alkaloid, is primarily sourced from endangered Dendrobium orchids, posing sustainability challenges to its production. Endophytic fungi, such as Trichoderma longibrachiatum MD33, offer an alternative; however, unresolved biosynthetic pathways and low yields hinder industrial scalability. Enhancing fungal metabolism through nanotechnology could address these limitations; however, nanoparticle-mediated engineering remains unexplored for dendrobine biosynthesis. This study aimed to (1) optimize dendrobine production in T. longibrachiatum MD33 using gold nanoparticles (CH-AuNPs) functionalized with alkaloid precursors and (2) elucidate the biosynthetic pathway to enable targeted metabolic engineering. CH-AuNPs were chemically synthesized, functionalized with L-phenylalanine, L-tyrosine, and tyramine, and applied to fungal cultures at concentrations of 0.5-20.0 mg/L. Multi-omics analyses (transcriptomics, proteomics, and metabolomics) identified pathway enzymes, and oxidative stress markers and dendrobine yields were quantified. Dose-dependent CH-AuNP exposure (10.0mg/L optimal) elevated dendrobine production by 63.7%, balancing pathway activation and oxidative stress. Multi-omics analysis revealed a hybrid terpenoid-alkaloid pathway, wherein sesquiterpene scaffolds from the mevalonate pathway merge with ornithine-derived piperidine moieties. This process is regulated by sesquiterpene synthases (TPS), cytochrome P450s (CYP71D1), and O-methyltransferases (COMT). Metabolomic analysis provided direct evidence for the rechanneling of nitrogen metabolism, with depletion of glutamate and ornithine pools and accumulation of polyamine pathway intermediates such as putrescine, supporting the transcriptional upregulation of ornithine decarboxylase (ODC). Mechanistically, low-to-moderate oxidative stress induced by CH-AuNPs activated redox-sensitive transcription factors and stress-responsive pathways, which in turn upregulated terpenoid and alkaloid biosynthesis genes. This controlled stress response enhanced precursor flux and enzyme activity, leading to increased dendrobine synthesis without triggering cellular damage in the cells. Concentrations > 10.0mg/L suppressed metabolism owing to oxidative damage. CH-AuNPs act as precision tools to upregulate dendrobine biosynthesis in T. longibrachiatum MD33, resolving the hybrid pathway and establishing this fungus as a sustainable production platform for dendrobine. The dose-dependent response highlights the dual role of nanoparticle-mediated engineering in metabolic enhancement and stress induction. This integration of nanotechnology and multi-omics bridges the critical gaps in fungal biotechnology, enabling scalable and eco-friendly alkaloid synthesis. Future applications include CRISPR-AuNP genome editing and bioreactor optimization, which will advance pharmaceutical and environmental biotechnologies.

Dendrobium belongs to one of the most important genera of the Orchidaceae family, which covers the largest category of flowering plants. More recent traditional medicinal studies have revealed that these orchid plants are used worldwide to treat a variety of illnesses, including those related to the complexion, pulmonary, gastrointestinal, reproductive, and circulatory systems, as well as cancers, pain, and inflammation. Several Dendrobium species have been the subject of phytochemistry and pharmacological research to explore their different medicinal aspects. A comprehensive search through databases (PubMed, Google Scholar, Web of Science, Traditional integrated knowledge resources, local dissertations, and books) was conducted up until November 2024 using key terms such as "Orchid plants, Type 2 diabetes, phytomedicine, natural products, phytoconstituents, phytochemistry, therapeutic potential, and plantderived antidiabetic agents." Its major phytoconstituents belong to the chemical categories of stilbenoid, glycoside, alkaloids, flavonoids, phenanthrenes, anthocyanins, carotenoids, and bibenzyl derivatives, in which the alkaloids and flavonoids play a most significant role for their biological properties. The flavone Cglycoside and flavanols are commonly found, but highly methylated and glycosylated derivatives of flavonoids are absent in orchid leaves. The phytochemical composition of Dendrobium, along with its structure-activity relationship of major classes, supports it as a potential source for novel antidiabetic agents through integrating traditional knowledge with medicinal chemistry aspects. A specific Dendrobium orchid plant has a promising role in the effective management of diabetes. In the future, newer, more potent antidiabetic compounds may be isolated and further developed into more effective antidiabetic agents.

Introduction: The limitation of endosperm in seeds requires the propagation of Dendrobium orchid plants to be carried out using in vitro culture techniques. The composition of in vitro culture media needs to be considered because it greatly determines the success of plant propagation. The addition of sucrose and myo-inositol to the growth media can support the growth of explants planted in vitro. This study aimed to determine the best concentration of myo-inositol and sucrose added to the multiplication medium to increase the growth of Dendrobium orchid explants. Methods: This study used a Completely Randomized Design. The experimental treatments were the concentration of myo-inositol and sucrose added to the VW media consisting of four levels, namely myo-inositol 50 mg/l + sucrose 20 mg/l, myo-inositol 50 mg/l + sucrose 40 mg/l, myo-inositol 100 mg/l + sucrose 20 mg/l and myo-inositol 100 mg/l + sucrose 40 mg/l. Observation variables include plant height, shoot emergence time, number of shoots and shoot growth rate. Data were analyzed using analysis of variance (ANOVA) at 5% α level and Duncan's Multiple Range Test (DMRT) at 5% level. Results: The addition of 40 g/l sucrose + 50 mg/l myo-inositol to the in vitro culture medium was the best treatment in producing optimal explant growth. Conclusion: The addition of 40 g/l sucrose can reduce the addition of myo-inositol to VW media by up to 50% to propagate Dendrobium orchids in vitro.

IntroductionDendrobium is a multi-purpose medicinal orchid that grows on the edge of high­altitude cliffs. The content of water-soluble polysaccharides (WSPs), which primarily play a pharmacological role, is the main criterion for evaluating the quality of Dendrobium orchid. Therefore, it is necessary to study the regulatory manner involved in the accumulation of WSPs.MethodsD. officinale were treated with methyl jasmonate (MeJA), and WSPs content was measured at different time points to assess the dynamic accumulation pattern. To clarify the role of one finger (Dof) family genes in the MeJA-mediated WSP metabolic pathway, a bioinformatics analysis identified Dof members in D. huoshanense, D. nobile and D. officinale. Based on expression patterns and co-expression analysis, a regulatory factor, DoDof4 was identified.ResultsIn this study, the elicitation of D. officinale by methyl jasmonate (MeJA) increased WSP production, which was further amplified by extending the treatment period. Analysis of transcriptomic data revealed that members of the DNA-binding with Dof gene family members accounted for 4% of all differentially expressed genes coding for transcription factors, following MeJA induction. To clarify the role of Dof family genes in the MeJA-mediated WSP metabolic pathway, a bioinformatics analysis identified 29, 29 and 22 Dof members in D. huoshanense, D. nobile and D. officinale, respectively and these were divided into four groups. DoDof4 was encoded a 31.16 kDa protein composed of 292 amino acids, and was targeted on chromosome 3. Furthermore, DoDof4 was a typical transcription factor that localized in the nucleus, displayed transcriptional activity and increased the WSPs accumulation. DoDof4 was co-expressed with 15 genes involved in the WSP metabolic pathway, eight of which displayed a positive Pearson's correlation coefficient. Additionally, correlation analysis revealed the possible downstream targets (KM980199 and KP203853) of DoDof4.DiscussionThe results of the present study suggest that DoDof4 acts as an important regulator in the WSPs metabolic pathway, exhibiting potential values for the improvement of WSPs in Dendrobium species.

This study investigated the occurrence of insects associated to the cultivation of orchid Dendrobium phalaenopsis Fitzg. in nurseries in Bahia State. The analyses were performed through Dominance indices (D), Shannon-Weiner diversity (H’), Margalef and Pielou’s Equitability (J’), while the Pearson correlation analysis was used to verify the correlation between the occurrence magnitude of insects with environmental variables (temperature, humidity, precipitation, and wind speed). Univariate statistics were used through a cross contingency table, differentiating classes from categories using the Chi-square test and the multiple correspondence analysis. We collected 927 adult individuals and the Coccinellidae (Coleoptera) family was the most abundant. We obtained 39 genera, distributed in 49 morphospecies, and most taxa identified were collected on the leaves of orchids. The most abundant taxa were Diomus spp. (Coleoptera: Coccinellidae), Leptophysa sp.1 (Coleoptera: Chrysomelidae), Thaliabaris sp. (Coleoptera: Curculionidae), Dynaspidiotus sp., Furcaspis sp.1 (Hemiptera: Diaspididae), Tenthecoris sp. (Hemiptera: Miridae), and Pheidole sp. (Hymenoptera: Formicidae). The large commercial nursery showed the highest species diversity with Shannon-Wiener index (H’=3.008), Margalef (6.454), and dominance (D=0.071). Species uniformity was higher in the small commercial nursery (J’=0.9579). This study contributes to the valuation of entomofauna of orchid and it is the first list of insects associated to orchid cultivation in Bahia State, Brazil. It also presents the first record of mealybug Dynaspidiotus sp. and beetles Leptophysa sp. and Thaliabaris sp. in orchid cultivation in Bahia State.

Abstract Orchid Dendrobium is an economically ornamental plant. However, the propagation of This Plant faces several obstacles, including the low production of high-quality seedlings. Somatic embryogenesis is a tissue culture technique currently considered the most appropriate to increase the production of high-quality orchid seedlings. Most previous studies have focused on the early induction of somatic embryogenesis under different media compositions. However, the success of somatic embryogenesis is also determined at the proliferation stage. Inaccuracy in media concentration and plant growth regulator determination can reduce the quality of propagation results. This study focused on the proliferation stage, which has the potential to determine the success of planlet quality. This study aimed to obtain the best combination of NAA and TDZ on the somatic proliferation of Dendrobium orchid through somatic embryogenesis. The results showed that the best combination of NAA and TDZ in the N2T3 medium treatment (0.2 mg/L NAA + 0.4 mg/L TDZ) gave the best response in forming globular with the fastest average time of 3 days, forming scutellar with the fastest average time of 6 days, and coleoptile with a mean of 20 days. These propagation protocols could benefit commercial production, especially for large-scale production of Dendrobium using liquid media.

This study investigates the phenotypic alterations in Dendrobium 'Transient White Rika' (RKA) and 'Florenza' (FLO) orchids resulting from colchicine-induced polyploidy during in vitro cultivation. Colchicine, a known mitotic inhibitor, was applied at varying concentrations (0 ppm, 4000 ppm, 5000 ppm, 6000 ppm) to induce chromosome doubling and assess its impact on the vegetative and generative phases of the orchids. A factorial Randomized Block Design (RBD) was employed to analyze the effects of these treatments. The study revealed significant variations in leaf length and color, particularly at higher colchicine concentrations. Morphological changes were most pronounced at 6000 ppm, where the leaf length decreased, and flower morphology exhibited unique features compared to the control. The study also observed a reduction in stomatal density as colchicine concentration increased, correlating with an enlargement of leaf epidermal cells. These findings indicate that colchicine treatment can effectively induce phenotypic changes in Dendrobium orchids, contributing to the development of hybrids with superior ornamental qualities. However, the study also notes that the response to colchicine is variable, with certain parameters like the number of new shoots and leaf width not showing significant changes. The results underscore the potential of colchicine-induced polyploidy in orchid breeding, though further research is necessary to optimize treatment protocols and ensure the stability and viability of the resulting hybrids. These findings may serve as valuable material for genetics-based learning modules in biology.

This study aims to obtain a good concentration of IBA and plantain and is suitable for root induction in Dendrobium sp. The benefit of this study is to provide information about the successful root induction of orchid plantlets Dendrobium sp. In vitro culture and creating formulations using IBA hormones and organic plantain ingredients in in vitro culture media optimally. This research has been carried out at the YAHDI Plant Tissue Culture Laboratory. Rooting orchids Dendrobium sp. In vitro using MS basic media which was added with IBA (Indole 3 Butyric Acid) 0 ppm, 2 ppm, 4 ppm, and 6 ppm, and added with plantain pulp with levels of 0 g/l, 100 g/l. and 200 g/l. This study was a factorial experiment with 2 factors compiled based on a 4 x 3 Complete Random Design (RAL) with 12 treatment combinations. The data obtained was analyzed for variance analysis (ANOVA). If there are results that have a real effect, it is followed by the DMRT (Duncan Multiple Range Test) test at the level of 5%. (1) The results of the study showed that the treatment of the variables of the number of roots, plant height, and root length was influenced by IBA. (2) The treatment of the variables of the number of buds, the number of leaves, the number of roots, the height of the plant, and the length of the roots is influenced by plantains. (3) The treatment of the variables on the number of roots, root length, and leaf width were influenced by the interaction of IBA and banana pulp.

This study aims to assess the effect of young coconut water concentration on the growth of Dendrobium orchids. The research was conducted for 4 months at Candi Orchid Production House, Semarang, with a one-factor completely randomized design (CRD) and four treatments of young coconut water concentration (0 mL/L, 100 mL/L, 150 mL/L, 200 mL/L), each with five replications. Parameters observed included number of shoots, root length, and plant height. Data were analyzed using ANOVA and Duncan's further test at the 5% level. The results showed that the application of young coconut water had no significant effect on the number of shoots and root length (p > 0.05), but had a significant effect on plant height (p = 0.003), especially at a concentration of 100 mL/L. The predominance of cytokinin hormones in coconut water likely plays a role in stimulating stem growth, while its effects on shoots and roots require interactions with other hormones. Although coconut water contains essential nutrients, its composition may not be fully suitable for the physiological needs of orchids in the seedling phase.

The critically endangered orchid Dendrobium officinale, valued for its medicinal properties, depends on specific seedling-associated mycorrhizal fungi (SAMF) for successful early-stage seedling development. However, conservation efforts are often hindered by difficulties in obtaining suitable SAMF, leading to poor seedling establishment in both natural and cultivated environments. In this study, we explored the growth-promoting effects of SAMF and evaluated the performances of synthetic fungal combinations. Our results demonstrated that mycorrhizal fungi, widely distributed across multiple habitats with high isolation frequencies, significantly promoted the growth of D. officinale, with specific fungi favoring different growth parameters. Tulasnella sp. TP-2 and TP-3 significantly improved stem diameter and plant height by 2.622 mm and 4.621 cm, while Tulasnella sp. TP-8 significantly increased tillering by a factor of 4.47. Additionally, Tulasnella sp. TP-11 and TP-13 markedly increased the number of new leaves (4.45) and new roots (2.688), respectively, identifying them as essential core OMFs for D. officinale seedlings. Contrary to expectations, synthetic fungal combinations composed of core orchid mycorrhizal fungi (core OMFs) did not exhibit synergistic growth-promoting effects. Instead, pronounced offset effects were observed, indicating that interactions between fungi may introduce competition or inhibition, limiting their collective ability to enhance plant growth. Our results confirmed that the core OMFs significantly promoted the growth of D. officinale seedlings. These core OMFs can serve as essential components in specialized microbial fertilizers for D. officinale, improving growth efficiency and yield, and supporting the sustainable development of the D. officinale industry.

This study aims to investigate how various concentrations of rice-washed water influence the growth of Dendrobium sp. in the post-acclimatization phase. A Completely Randomized Design (CRD) with one factor and four treatments was used: P0 (0% rice-washed water), P1 (30% rice-washed water), P2 (40% rice-washed water), and P3 (50% rice-washed water). The statistical analysis was conducted using a One-Way ANOVA at a 5% level. If significant differences were found, further analysis was performed using Duncan's Multiple Range Test. The results of data analysis revealed that (1) There were no significant differences in plant height and root length of Dendrobium orchids across the treatments during the post-acclimatization phase; (2) Treatments P1 to P3 did not yield optimal results in terms of plant height, as these concentrations were not suitable for supporting the height growth of Dendrobium orchids in the post-acclimatization phase; (3) The P2 concentration resulted in the longest root growth at 2.39 cm, while the shortest root length was observed in the P0 concentration. Thus, although the different concentrations of rice washing water did not produce statistically significant differences, the P2 concentration showed potential for promoting root growth in Dendrobium orchids during the post-acclimatization phase.

Nitric oxide has been identified as a chemical compound that can prolong the vase life of various cut flowers. The effects of sodium nitroprusside, a nitric oxide donor, on 1-aminocyclopropane-1-carboxylate synthase and 1-aminocyclopropane-1-carboxylic Acid oxidase activities, ethylene production, respiration, bud opening, and electrolyte leakage and lipid peroxidation during post-harvest storage of cut inflorescences of Dendrobium cv. “Planty Fushia” orchid were evaluated. Inflorescences kept in 10 μmol L-1 sodium nitroprusside solution at room temperature (23±2°C) had longer vase life compared to controls in distilled water. Sodium nitroprusside increased vase life due to 1-aminocyclopropane-1-carboxylate synthase and 1-aminocyclo-propane-1-carboxylic acid oxidase a activities in buds and open flowers, thereby reducing ethylene production. Electrolyte leakage, malondialdehyde and respiration were also lower in plants treated with sodium nitroprusside compared to controls. In addition, sodium nitroprusside treatment increased the percentage of buds that opened. Therefore, sodium nitroprusside can be used to prolong the vase life of cut inflorescences of Dendrobium for 4 days at room temperature. The results indicate that sodium nitroprusside solution significantly affected the vase life of cut inflorescences of Dendrobium, 10 μmol L-1 sodium nitroprusside could significantly increase bud opening and prolong vase life compared to the control.

Eria hyacinthoides (Blume) Lindl. is an Indonesian orchid species found in Sumatra, Java, and Bali. This orchid is a sympodial orchid with flowers that has fragrant aroma, suspected containing phytochemicals for herbal medicines, so mass plant propagation is necessary. The aim of this research is to obtain the best in vitro conditions for this orchid through somatic embryos using growth regulators and analysing the structure of the Dendrobium Orchid Homeobox 1 (DOH1) homologous gene in E. hyacinthoides to Dendrobium ‘Madame Thong-In’ which is known to induce bud formation. The method used in this study: (1) the leaves of the plant spread about 20 - 30 days from shoots measuring about 6.3-6.7cm on the mother plant aged ± 8 years, stored in an incubation room with picture of 16 hours of light and 8 hours of darkness in the heat. 25 ± 1 ℃, (2) compared Murashige and Skoog, Vacin and Went, Knudson C and New Phalaenopsis growth to get the best medium, (3) added PGR to medium (Thidiazuron (TDZ) 1 - 3 mg L-1 and Naphthalene-1-acetic acid (NAA) 1-3 mg L-1), (4) isolate partial gen DOH1 homologous by using primer of DOH1, (5) analyse sequence of PCR products. Optimal medium for callus embryogenesis production from leave was Knudson C + TDZ 1 mg L-1 + NAA 1 mg L-1. Amplification of DNA fragments using degenerate primers of DOH1 resulted 175 bp, indicating similarity about 88.64 % with between the DOH1 gene structure in E. hyacinthoides and in Dendrobium ‘Madame Thong-In’.

Dendrobine, a significant medicinal compound, typically accumulates at low concentrations within several Dendrobium species, including Dendrobium nobile, Dendrobium catenatum, and Dendrobium moniliforme. As D. nobile and D. catenatum are established ingredients in traditional Chinese medicine and have been cultivated extensively, they present ideal plant chassis for upscaling dendrobine production for industrial and research applications. This study employed two approaches: the ectopic overexpression of seven genes through multigene stacking and the activation of multiple key endogenous genes in the dendrobine synthesis pathway using CRISPR/Act3.0 in either D. nobile or D. catenatum. These methods enhanced dendrobine production in transiently infiltrated leaves by 30.1% and transgenic plants by 35.6%. The study is the first to apply CRISPR/Act3.0 to Dendrobium orchids, enhancing dendrobine production, and thus laying a solid foundation for further improvements. CRISPR/Act3.0 is a recently developed technique that demonstrates high efficiency in model plant species, including rice, maize, and Arabidopsis. By combining CRISPR with transcriptional regulatory modules, activation of multiple endogenous genes in the metabolic pathway can be achieved. The successful application in orchid molecular breeding reveals promising potential for further exploration.

Comparative genome analysis unravels adaptive evolution in Dendrobium orchids.

Drought and low rainfall are natural phenomena that occurred in 2022-2023. These factors (abiotic factors) caused diseases in orchids, which affect the metabolism of growth and the flowering process, making the plants unable to survive in their habitat because they cannot reproduce, thus classifying them under Appendix 2 (CITES). However, some orchid species that are endemic are able to withstand drought and low rainfall. It is suspected that the orchids' ability to endure these factors is due to the role of endophytic fungi (Rhizoctonia binucleate / BNR), which can associate with orchids and produce secondary metabolites that help supply nutrients to the orchids during drought stress. This study aims to evaluate the biological resistance of different strains of Rhizoctonia binucleate / BNR isolates in Dendrobium orchids to understand their impact on water stress. The research method used a completely randomized design with a single stage consisting of two factors and five replications. Factor 1: orchid seedling types consisting of S1 = Phalaenopsis violaceae (epiphytic orchid) and S2 = Dendrobium aggregatum (epiphytic orchid). Factor 2: types of Rhizoctonia mycorrhiza application consisting of K0 = control, K1 = Rhizoctonia mycorrhiza application.

To assess the genetic variability, heritability, and association of 21 key quantitative variables in 15 distinct Dendrobium species, efforts were made to improve the germplasm of Dendrobium orchids in different parts of Arunachal Pradesh between 2020 and 2022. Significant variance was found in every attribute in the study, indicating that there is sufficient genetic diversity to support breeding efforts. The most significant values of PCV and GCV were recorded for the number of flowers per inflorescence (136.13% and 135.35%, respectively) and the number of inflorescences per plant (136.18% and 133.94%, respectively). High heritability coupled with high genetic advance was observed for inflorescence length (99.74%), the number of flowers per inflorescence (99.68%), flower width in the front view (99.41%), and flower length (99.25%). Selection based on these characteristics would be more effective for improving interspecific hybrids. Other characters exhibited moderate broad-sense heritability with low genetic advance. Plant height, internode number, number of leaves, and flower longevity showed positive and highly significant associations with the number of flowers per inflorescence and the number of inflorescences per plant at both the genotypic and phenotypic levels. However, correlation study revealed that selecting parents based on plant height, internode number, number of leaves, and flower longevity in plants would be helpful in breeding programs.

Effect of method of application and different nutrient solutions in Dendrobium orchid under polyhouse

The endangered orchid Dendrobium nobile is the primary source of dendrobine, a neuroprotective sesquiterpene alkaloid, but unsustainable harvesting necessitates alternative production platforms, such as the endophytic fungus Trichoderma longibrachiatum MD33. However, the fungal dendrobine pathway and its regulatory mechanisms remain uncharacterized, limiting its biotechnological exploitation. This study investigated cinnabar (HgS)-induced hormesis to elucidate the stress-mediated metabolic reprogramming of dendrobine biosynthesis through integrated transcriptomic and metabolomic profiling. Subtoxic HgS concentrations (1.0-4.0 μg/L) triggered ROS signaling, upregulating the mevalonate pathway, terpenoid synthases (TPS1/2), and cytochrome P450 monooxygenases (CYP450s), thereby enhancing dendrobine yields by 24% at 4.0 μg/L. In contrast, cytotoxic doses (6.0 μg/L) suppressed growth (73.9% inhibition) and dendrobine synthesis (73.2% reduction), correlating with metabolic collapse via disrupted CoA biosynthesis and antioxidant depletion. Multi-omics integration revealed biphasic regulation: low HgS activated stress-responsive transcription factors (bZIP, Zn-Cys6) and MAP kinase cascades, redirecting resources toward dendrobine production, whereas high HgS induced apoptotic markers and oxidative damage. These findings establish 4.0 μg/L a hormetic threshold for maximizing dendrobine yields and delineating the genetic and enzymatic architecture of its fungal biosynthesis. This work provides a roadmap for the metabolic engineering of T. longibrachiatum MD33, emphasizing ROS-mediated pathway optimization for sustainable alkaloid production. Future studies should leverage CRISPR-based editing of identified regulatory nodes (e.g., HMGR and FPPS) to enhance stress resilience and dendrobine titers in industrial strains.

Vanda is a monopodial epiphyte orchid that spreads throughout Asia and Southeast Asia reaching 70 species. Indonesia itself has its own endemic Vanda orchid such as Vanda tricolor and Vanda limbata. A hybrid of V. tricolor and V. limbata is predicted to form a new specific character in the flower and leaf. The purpose of this study was to determine the morphological and molecular differences between V. tricolor, V. limbata, and Vanda hybrids resulting from crosses between that two orchids, by analysing the morphological characteristics of the roots, leaves, flowers and the structure of the Vanda Orchid Homeobox1 (VOH1) shoot‐forming gene isolated from V. tricolor, V. limbata, and their hybrids. The morphological analysis was conducted using RHS colour chart, size measurement of plants, and the transversal preparation of the leaf. Molecular analysis was performed by PCR using Dendrobium Orchid Homeobox 1 (DOH1) primers, followed by sequencing and bioinformatic analysis. Morphologically, the flower’s colour of the hybrid is most similar to V. limbata but the flower’s patterns are more similar to V. tricolor meanwhile the leaf colour of the hybrid is brighter than the both parents. The slides illustrate the sclerenchyma tissue is made up of strongly thickened walls containing lignin indicates the presence of homeobox DOH1 gene homolog, namely VOH1. The molecular result displayed by the phylogenetic tree of the VOH1 indicates that the hybrid has more similarities with V. limbata.