Genome-Wide Survey Indicates Diverse Physiological Roles of Dendrobium officinale Calcium-Dependent Protein Kinase Genes.

Calcium-dependent protein kinases (CDPKs) are crucial calcium ions (Ca2+) sensors in plants with important roles in signal transduction, plant growth, development, and stress responses. Here, we identified 24 genes encoding CDPKs in Dendrobium officinale using genome-wide analysis. The phylogenetic analysis revealed that these genes formed four groups, with similar structures in the same group. The gene expression patterns following hormone treatments and yeast two-hybrid of homologous CDPK gene pairs with Rbohs showed differences, indicating functional divergence between homologous genes. In addition, the rapid accumulation of hydrogen peroxide (H2O2) and stomatal closure was observed in response to salicylic acid (SA)/jasmonic acid (JA) stress. Our data showed that CDPK9-2 and CDPK20-4 interacted with Rboh D and Rboh H, respectively, and were implicated in the generation of H2O2 and regulation of the stomatal aperture in response to salicylic acid/jasmonic acid treatment. We believe these results can provide a foundation for the functional divergence of homologous genes in D. officinale.

Transcriptome and Metabolome Reveal Salt-Stress Responses of Leaf Tissues from Dendrobium officinale.

Dendrobium officinale Kimura et Migo is a precious traditional Chinese medicine. Despite D. officinale displaying a good salt-tolerance level, the yield and growth of D. officinale were impaired drastically by the increasing soil secondary salinization. The molecular mechanisms of D. officinale plants' adaptation to salt stress are not well documented. Therefore, in the present study, D. officinale plants were treated with 250 mM NaCl. Transcriptome analysis showed that salt stress significantly altered various metabolic pathways, including phenylalanine metabolism, flavonoid biosynthesis, and α-linolenic acid metabolism, and significantly upregulated the mRNA expression levels of DoAOC, DoAOS, DoLOX2S, DoMFP, and DoOPR involved in the jasmonic acid (JA) biosynthesis pathway, as well as rutin synthesis genes involved in the flavonoid synthesis pathway. In addition, metabolomics analysis showed that salt stress induced the accumulation of some compounds in D. officinale leaves, especially flavonoids, sugars, and alkaloids, which may play an important role in salt-stress responses of leaf tissues from D. officinale. Moreover, salt stress could trigger JA biosynthesis, and JA may act as a signal molecule that promotes flavonoid biosynthesis in D. officinale leaves. To sum up, D. officinale plants adapted to salt stress by enhancing the biosynthesis of secondary metabolites.

Cytological Observation and Transcriptome Comparative Analysis of Self-Pollination and Cross-Pollination in Dendrobium Officinale.

Dendrobium officinale is a rare and traditional medicinal plant with high pharmacological and nutritional value. The self-incompatibility mechanism of D. officinale reproductive isolation was formed in the long-term evolution process, but intraspecific hybridization of different germplasm resources leads to a large gap in the yield, quality, and medicinal value of D. officinale. To investigate the biological mechanism of self-incompatibility in D. officinale, cytological observation and the transcriptome analysis was carried out on the samples of self-pollination and cross-pollination in D. officinale. Results for self-pollination showed that the pollen tubes could grow in the style at 2 h, but most of pollen tubes stopped growing at 4 h, while a large number of cross-pollinated pollen tubes grew along the placental space to the base of ovary, indicating that the self-incompatibility of D. officinale may be gametophyte self-incompatibility. A total of 63.41 G basesum of D. officinale style samples from non-pollinated, self-pollination, and cross-pollination by RNA-seq were obtained, and a total of 1944, 1758, and 475 differentially expressed genes (DEGs) in the comparison of CK (non-pollinated) vs. HF (cross-pollination sample), CK vs. SF (self-pollination sample) and SF vs. HF were identified, respectively. Forty-one candidate genes related to self-incompatibility were found by function annotation of DEGs, including 6 Ca2+ signal genes, 4 armed repeat containing (ARC) related genes, 11 S-locus receptor kinase (SRK) related genes, 2 Exo70 family genes, 9 ubiquitin related genes, 1 fatty acid related gene, 6 amino acid-related genes, 1 pollen-specific leucine-rich repeat extensin-like protein (LRX) related gene and 1 lectin receptor-like kinases (RLKs) related gene, showed that self-incompatibility mechanism of D. officinale involves the interaction of multiple genes and pathways. The results can provide a basis for the study of the self-incompatibility mechanism of D. officinale, and provide ideas for the preservation and utilization of high-quality resources of D. officinale.

The water-water cycle is not a major alternative sink in fluctuating light at chilling temperature.

The water-water cycle (WWC) has the potential to alleviate photoinhibition of photosystem I (PSI) in fluctuating light (FL) at room temperature and moderate heat stress. However, it is unclear whether WWC can function as a safety valve for PSI in FL at chilling temperature. In this study, we measured P700 redox state and chlorophyll fluorescence in FL at 25 °C and 4 °C in the high WWC activity plant Dendrobium officinale. At 25 °C, the operation of WWC contributed to the rapid re-oxidation of P700 upon dark-to-light transition. However, such rapid re-oxidation of P700 was not observed at 4 °C. Upon a sudden increase in light intensity, WWC rapidly consumed excess electrons in PSI and thus avoided an over-reduction of PSI at 25 °C. On the contrary, PSI was highly reduced within the first seconds after transition from low to high light at 4 °C. Therefore, in opposite to 25 °C, the WWC is not a major alternative sink in FL at chilling temperature. Upon transition from low to high light, cyclic electron transport was highly stimulated at 4 °C when compared with 25 °C. These results indicate that D. officinale enhances cyclic electron transport to partially compensate for the inactivation of WWC in FL at 4 °C.

Photosystem I is tolerant to fluctuating light under moderate heat stress in two orchids Dendrobium officinale and Bletilla striata.

Under natural field conditions, plants usually experience fluctuating light (FL) under moderate heat stress in summer. However, responses of photosystems I and II (PSI and PSII) to such combined stresses are not well known. Furthermore, the role of water-water cycle (WWC) in photoprotection in FL under moderate heat stress is poorly understood. In this study, we examined chlorophyll fluorescence and P700 redox state in FL at 42 °C in two orchids, Dendrobium officinale (with high WWC activity) and Bletilla striata (with low WWC activity). After FL treatment at 42 °C, PSI activity maintained stable while PSII activity decreased significantly in these two orchids. In D. officinale, the WWC could rapidly consume the excess excitation energy in PSI and thus avoided an over-reduction of PSI upon any increase in illumination. Therefore, in D. officinale, WWC likely protected PSI in FL at 42 °C. In B. striata, heat-induced PSII photoinhibition down-regulated electron flow from PSII and thus prevented an over-reduction of PSI after transition from low to high light. Consequently, in B. striata moderate PSII photoinhibition could protected PSI in FL at 42 °C. We conclude that, in addition to cyclic electron flow, WWC and PSII photoinhibition-repair cycle are two important strategies for preventing PSI photoinhibition in FL under moderate heat stress.

A perspective on crassulacean acid metabolism photosynthesis evolution of orchids on different continents: Dendrobium as a case study.

Members of the Orchidaceae, one of the largest families of flowering plants, evolved the crassulacean acid metabolism (CAM) photosynthesis strategy. It is thought that CAM triggers adaptive radiation into new niche spaces, yet very little is known about its origin and diversification on different continents. Here, we assess the prevalence of CAM in Dendrobium, which is one of the largest genera of flowering plants and found in a wide range of environments, from the high altitudes of the Himalayas to relatively arid habitats in Australia. Based on phylogenetic time trees, we estimated that CAM, as determined by δ 13C values less negative than -20.0‰, evolved independently at least eight times in Dendrobium. The oldest lineage appeared in the Asian clade during the middle Miocene, indicating the origin of CAM was associated with a pronounced climatic cooling that followed a period of aridity. Divergence of the four CAM lineages in the Asian clade appeared to be earlier than divergence of those in the Australasian clade. However, CAM species in the Asian clade are much less diverse (25.6%) than those in the Australasian clade (57.9%). These findings shed new light on CAM evolutionary history and the aridity levels of the paleoclimate on different continents.

Comparative nutritional characteristics of the three major Chinese Dendrobium species with different growth years.

Dendrobium, an important medicinal plant, is a source of widely used herbal medicine to nourish the stomach and treat throat inflammation. The present study is aimed at distinguishing and evaluating three major Dendrobium species by comparing physiochemical characteristics and understanding differences between different growth years in the Ta-pieh Mountains. Polysaccharides and total alkaloids of Dendrobium were determined, and the amino acids and trace elements were determined by UPLC (Ultra High-Performance Liquid Chromatography) and ICP-MS (Inductively coupled plasma mass spectrometry). It can be seen from the results that the polysaccharide content of these three kinds of Dendrobium in different growth years ranges from 249.31 mg·g-1 to 547.66 mg·g-1, and the highest content is in the 3-year-old Dendrobium huoshanense. The total alkaloid content ranges from 0.21 mg·g-1 to 0.54 mg·g-1, and the highest content is also the 3-year-old Dendrobium huoshanense. We determined the amino acid content of these three Dendrobium in different growth years, and we can see that each of the three kinds of Dendrobium contain seven kinds of amino acids required by the human body. We conducted a safety evaluation of the essential trace elements of Dendrobium, and the results showed that the dosage of 12g·d-1 Dendrobium prescribed in China Pharmacopoeia is in accordance with the recommended daily intake of trace elements recommended by the Food and Drug Administration of the United States, and will not cause trace element poisoning. Linear discriminant analysis was carried out on the basis of amino acids and trace elements and confirmed the applicability of multi-elemental analysis for identifying different Dendrobium species.

Circadian Regulation of Alternative Splicing of Drought-Associated CIPK Genes in Dendrobium catenatum (Orchidaceae).

Dendrobium catenatum, an epiphytic and lithophytic species, suffers frequently from perennial shortage of water in the wild. The molecular mechanisms of this orchid's tolerance to abiotic stress, especially drought, remain largely unknown. It is well-known that CBL-interacting protein kinase (CIPKs) proteins play important roles in plant developmental processes, signal transduction, and responses to abiotic stress. To study the CIPKs' functions for D. catenatum, we first identified 24 CIPK genes from it. We divided them into three subgroups, with varying intron numbers and protein motifs, based on phylogeny analysis. Expression patterns of CIPK family genes in different tissues and in response to either drought or cold stresses suggested DcaCIPK11 may be associated with signal transduction and energy metabolism. DcaCIPK9, -14, and -16 are predicted to play critical roles during drought treatment specifically. Furthermore, transcript expression abundances of DcaCIPK16 showed polar opposites during day and night. Whether under drought treatment or not, DcaCIPK16 tended to emphatically express transcript1 during the day and transcript3 at night. This implied that expression of the transcripts might be regulated by circadian rhythm. qRT-PCR analysis also indicated that DcaCIPK3, -8, and -20 were strongly influenced by circadian rhythmicity. In contrast with previous studies, for the first time to our knowledge, our study revealed that the major CIPK gene transcript expressed was not always the same and was affected by the biological clock, providing a different perspective on alternative splicing preference.

Functional Insights into the Roles of Hormones in the Dendrobium officinale-Tulasnella sp. Germinated Seed Symbiotic Association.

Dendrobium is one of the largest genera in the Orchidaceae, and D. officinale is used in traditional medicine, particularly in China. D. officinale seeds are minute and contain limited energy reserves, and colonization by a compatible fungus is essential for germination under natural conditions. When the orchid mycorrhizal fungi (OMF) initiates symbiotic interactions with germination-driven orchid seeds, phytohormones from the orchid or the fungus play key roles, but the details of the possible biochemical pathways are still poorly understood. In the present study, we established a symbiotic system between D. officinale and Tulasnella sp. for seed germination. RNA-Seq was used to construct libraries of symbiotic-germinated seeds (DoTc), asymbiotic-germinated seeds (Do), and free-living OMF (Tc) to investigate the expression profiles of biosynthesis and metabolism pathway genes for three classes of endogenous hormones: JA (jasmonic acid), ABA (abscisic acid) and SLs (strigolactones), in D. officinale seeds and OMF under symbiotic and asymbiotic conditions. Low concentrations of endogenous JA, ABA, or SLs were detected in the D. officinale-Tulasnella symbiont compared with the asymbiotic tissues. Gene annotation results suggest that the expression of DEGs (differentially expressed genes) related to JA and ABA biosynthesis from D. officinale were down-regulated, while most of the key DEGs related to SL biosynthesis from D. officinale were up-regulated in the symbiotic germinated seeds compared with the asymbiotic germinated seeds. Moreover, in the OMF, we found a significantly up-regulated differential expression of the JA and ABA biosynthesis-related genes in the symbiotic interaction, with the opposite expression trends to those found in Dendrobium. This indicates that Dendrobium seed symbiotic germination may be stimulated by the apparent involvement of the OMF in the production of hormones, and relatively low concentrations of endogenous JA, ABA, or SLs might be maintained to promote the growth of the D. officinale-Tulasnella symbiotic protocorm-like body. These results will increase our understanding of the possible roles played by endogenous hormones in the regulation of the orchid-fungus symbiosis.

Genome-wide researches and applications on Dendrobium.

MAIN CONCLUSION: This review summarizes current knowledge of chromosome characterization, genetic mapping, genomic sequencing, quality formation, floral transition, propagation, and identification in Dendrobium. The widely distributed Dendrobium has been studied for a long history, due to its important economic values in both medicine and ornamental. In recent years, some species of Dendrobium and other orchids had been reported on genomic sequences, using the next-generation sequencing technology. And the chloroplast genomes of many Dendrobium species were also revealed. The chromosomes of most Dendrobium species belong to mini-chromosomes, and showed 2n = 38. Only a few of genetic studies were reported in Dendrobium. After revealing of genomic sequences, the techniques of transcriptomics, proteomics and metabolomics could be employed on Dendrobium easily. Some other molecular biological techniques, such as gene cloning, gene editing, genetic transformation and molecular marker developing, had also been applied on the basic research of Dendrobium, successively. As medicinal plants, insights into the biosynthesis of some medicinal components were the most important. As ornamental plants, regulation of flower related characteristics was the most important. More, knowledge of growth and development, environmental interaction, evolutionary analysis, breeding of new cultivars, propagation, and identification of species and herbs were also required for commercial usage. All of these studies were improved using genomic sequences and related technologies. To answer some key scientific issues in Dendrobium, quality formation, flowering, self-incompatibility and seed germination would be the focus of future research. And genome related technologies and studies would be helpful.

Comparative Proteomics Analyses of Pollination Response in Endangered Orchid Species Dendrobium Chrysanthum.

Pollination is a crucial stage in plant reproductive process. The self-compatibility (SC) and self-incompatibility (SI) mechanisms determined the plant genetic diversity and species survival. D. chrysanthum is a highly valued ornamental and traditional herbal orchid in Asia but has been declared endangered. The sexual reproduction in D. chrysanthum relies on the compatibility of pollination. To provide a better understanding of the mechanism of pollination, the differentially expressed proteins (DEP) between the self-pollination (SP) and cross-pollination (CP) pistil of D. chrysanthum were investigated using proteomic approaches-two-dimensional electrophoresis (2-DE) coupled with tandem mass spectrometry technique. A total of 54 DEP spots were identified in the two-dimensional electrophoresis (2-DE) maps between the SP and CP. Gene ontology analysis revealed an array of proteins belonging to following different functional categories: metabolic process (8.94%), response to stimulus (5.69%), biosynthetic process (4.07%), protein folding (3.25%) and transport (3.25%). Identification of these DEPs at the early response stage of pollination will hopefully provide new insights in the mechanism of pollination response and help for the conservation of the orchid species.

DOFT and DOFTIP1 affect reproductive development in the orchid Dendrobium Chao Praya Smile.

FLOWERING LOCUS T (FT) in Arabidopsis encodes the florigen that moves from leaves to the shoot apical meristem to induce flowering, and this is partly mediated by FT-INTERACTING PROTEIN 1 (FTIP1). Although FT orthologs have been identified in some flowering plants, their endogenous roles in Orchidaceae, which is one of the largest families of flowering plants, are still largely unknown. In this study, we show that DOFT and DOFTIP1, the orchid orthologs of FT and FTIP1, respectively, play important roles in promoting flowering in the orchid Dendrobium Chao Praya Smile. Expression of DOFT and DOFTIP1 increases in whole plantlets during the transition from vegetative to reproductive development. Both transcripts are present in significant levels in reproductive organs, including inflorescence apices, stems, floral buds, and open flowers. Through successful generation of transgenic orchids, we have revealed that overexpression or down-regulation of DOFT accelerates or delays flowering, respectively, while alteration of DOFT expression also greatly affects pseudobulb formation and flower development. In common with their counterparts in Arabidopsis and rice, DOFTIP1 interacts with DOFT and affects flowering time in orchids. Our results suggest that while DOFT and DOFTIP1 play evolutionarily conserved roles in promoting flowering, DOFT may have evolved with hitherto unknown functions pertaining to the regulation of storage organs and flower development in the Orchidaceae family.

Influence of host tree species on isolation and communities of mycorrhizal and endophytic fungi from roots of a tropical epiphytic orchid, Dendrobium sinense (Orchidaceae).

Most studies on the host preference of orchids have focused on the association between orchids and host characteristics, but little is known about the differences of mycorrhizal and endophytic fungal communities in epiphytic orchids growing on different host tree species. We selected Dendrobium sinense, a tropical epiphytic orchid, to determine if fungal endophytes from the roots of D. sinense were preferentially correlated with host tree species. Fifty-six fungal operational taxonomic units (OTUs) from 36 host trees were identified. The results indicated that the species richness and diversity of mycorrhizal and endophytic fungal communities isolated from D. sinense roots were strongly influenced by host tree species. Both species richness and diversity indices showed that D. sinense roots on Syzygium buxifolium harbored the most diverse and abundant endophytic fungi. Species of Tulasnellaceae were dominant on S. buxifolium and Rhododendron moulmainense but infrequent on Cyclobalanopsis disciformis and Podocarpus neriifolius. Our results provide evidence for distinct mycorrhizal and endophytic fungal communities on different host tree species. Further research focusing on fungi-orchid-host preference could be conducted to increase our understanding for the in situ conservation of epiphytic orchids.

iTRAQ and RNA-Seq Analyses Provide New Insights into Regulation Mechanism of Symbiotic Germination of Dendrobium officinale Seeds (Orchidaceae).

Mycorrhizal fungi colonize orchid seeds and induce germination. This so-called symbiotic germination is a critical developmental process in the lifecycle of all orchid species. However, the molecular changes that occur during orchid seed symbiotic germination remain largely unknown. To better understand the molecular mechanism of orchid seed germination, we performed a comparative transcriptomic and proteomic analysis of the Chinese traditional medicinal orchid Dendrobium officinale to explore the change in protein expression at the different developmental stages during asymbiotic and symbiotic germination and identify the key proteins that regulate the symbiotic germination of orchid seeds. Among 2256 identified plant proteins, 308 were differentially expressed across three developmental stages during asymbiotic and symbiotic germination, and 229 were differentially expressed during symbiotic germination compared to asymbiotic development. Of these, 32 proteins were coup-regulated at both the proteomic and transcriptomic levels during symbiotic germination compared to asymbiotic germination. Our results suggest that symbiotic germination of D. officinale seeds shares a common signaling pathway with asymbiotic germination during the early germination stage. However, compared to asymbiotic germination, fungal colonization of orchid seeds appears to induce higher and earlier expression of some key proteins involved in lipid and carbohydrate metabolism and thus improves the efficiency of utilization of stored substances present in the embryo. This study provides new insight into the molecular basis of orchid seed germination.

ROS-INDUCED OXIDATIVE STRESS IN NOBILE-TYPE DENDROBIUM PROTOCORM-LIKE BODIES (PLBS) DURING VITRIFICATION.

BACKGROUND: Oxidative stress involved in cryopreservation protocols may be responsible for the poor survival of tissues after cryopreservation. OBJECTIVE: In the current study, we aimed to clarify the role of oxidative stress and its relationship with survival rate during cryopreservation of PLBs from nobile-type Dendrobium. MATERIALS AND METHODS: ROS, antioxidants and oxidative products and the survival rate in PLBs from Dendrobium Hamanal Lake Dream were determined during vitrification. RESULTS: Relative survival of PLBs decreased significantly after preculture and rewarming (P<0.01). Generation of ·O2- and protein carbonyl (PCO) increased significantly after preculture. Dramatic increases in ·O2-, H2O2 and MDA, and significant decreases in AsA content, activities of SOD and CAT were observed after rewarming. CONCLUSION: ROS-induced oxidative stress was associated with the poor survival of PLBs following vitrification. ·O2- was the predominant ROS resulting in the decreased survival after preculture, while H2O2 together with ·O2- appear to be responsible for the survival decrease after rewarming.

Cytotoxic and Antifungal Constituents Isolated from the Metabolites of Endophytic Fungus DO14 from Dendrobium officinale.

Two novel cytotoxic and antifungal constituents, (4S,6S)-6-[(1S,2R)-1, 2-dihydroxybutyl]-4-hydroxy-4-methoxytetrahydro-2H-pyran-2-one (1), (6S,2E)-6-hydroxy-3-methoxy-5-oxodec-2-enoic acid (2), together with three known compounds, LL-P880γ (3), LL-P880α (4), and Ergosta-5,7,22-trien-3b-ol (5) were isolated from the metabolites of endophytic fungi from Dendrobium officinale. The chemical structures were determined based on spectroscopic methods. All the isolated compounds 1-5 were evaluated by cytotoxicity and antifungal effects. Our present results indicated that compounds 1-4 showed notable anti-fungal activities (minimal inhibitory concentration (MIC) ≤ 50 µg/mL) for all the tested pathogens including Candida albicans, Cryptococcus neoformans, Trichophyton rubrum, Aspergillus fumigatus. In addition, compounds 1-4 possessed notable cytotoxcities against human cancer cell lines of HL-60 cells with the IC50 values of below 100 µM. Besides, compounds 1, 2, 4 and 5 showed strong cytotoxities on the LOVO cell line with the IC50 values were lower than 100 µM. In conclusion, our study suggested that endophytic fungi of D. officinale are great potential resources to discover novel agents for preventing or treating pathogens and tumors.

Vesicles between plasma membrane and cell wall prior to visible senescence of Iris and Dendrobium flowers.

Cut Iris flowers (Iris x hollandica, cv. Blue Magic) show visible senescence about two days after full opening. Epidermal cells of the outer tepals collapse due to programmed cell death (PCD). Transmission electron microscopy (TEM) showed irregular swelling of the cell walls, starting prior to cell collapse. Compared to cells in flowers that had just opened, wall thickness increased up to tenfold prior to cell death. Fibrils were visible in the swollen walls. After cell death very little of the cell wall remained. Prior to and during visible wall swelling, vesicles (paramural bodies) were observed between the plasma membrane and the cell walls. The vesicles were also found in groups and were accompanied by amorphous substance. They usually showed a single membrane, and had a variety of diameters and electron densities. Cut Dendrobium hybrid cv. Lucky Duan flowers exhibited visible senescence about 14 days after full flower opening. Paramural bodies were also found in Dendrobium tepal epidermis and mesophyll cells, related to wall swelling and degradation. Although alternative explanations are well possible, it is hypothesized that paramural bodies carry enzymes involved in cell wall breakdown. The literature has not yet reported such bodies in association with senescence/PCD.

dsRNA silencing of an R2R3-MYB transcription factor affects flower cell shape in a Dendrobium hybrid.

BACKGROUND: The R2R3-MYB genes regulate pigmentation and morphogenesis of flowers, including flower and cell shape, and therefore have importance in the development of new varieties of orchids. However, new variety development is limited by the long breeding time required in orchids. In this study, we identified a cDNA, DhMYB1, that is expressed during flower development in a hybrid orchid, Dendrobium hybrida (Dendrobium bobby messina X Dendrobium chao phraya) then used the direct application of dsRNA to observe the effect of gene silencing on flower phenotype and floral epidermal cell shape. RESULTS: Flower bud development in the Dendrobium hybrid was characterised into seven stages and the time of meiosis was determined as between stages 3 to 5 when the bud is approximately half of the mature size. Scanning electron microscopy characterisation of adaxial epidermal cells of the flower perianth, showed that the petals and sepals each are divided into two distinct domains based on cell shape and size, while the labellum comprises seven domains. Thirty-two partial cDNA fragments representing R2R3-MYB gene sequences were isolated from D. hybrida. Phylogenetic analysis revealed that nine of the translated sequences were clustered with MYB sequences that are known to be involved in cell shape development and from these, DhMYB1 was selected for full length cDNA cloning and functional study. Direct application of a 430 bp dsRNA from the 3' region of DhMYB1 to emerging orchid flower buds reduced expression of DhMYB1 RNA compared with untreated control. Scanning electron microscopy of adaxial epidermal cells within domain one of the labellum of flowers treated with DhMYB1 dsRNA showed flattened epidermal cells whilst those of control flowers were conical. CONCLUSIONS: DhMYB1 is expressed throughout flower bud development and is involved in the development of the conical cell shape of the epidermal cells of the Dendrobium hybrida flower labellum. The direct application of dsRNA changed the phenotype of floral cells, thus, this technique may have application in floriculture biotechnology.

Asymbiotic in vitro seed propagation of Dendrobium.

The ability to germinate orchids from seeds in vitro presents a useful and viable method for the propagation of valuable germplasm, maintaining the genetic heterogeneity inherent in seeds. Given the ornamental and medicinal importance of many species within the genus Dendrobium, this review explores in vitro techniques for their asymbiotic seed germination. The influence of abiotic factors (such as temperature and light), methods of sterilization, composition of basal media, and supplementation with organic additives and plant growth regulators are discussed in context to achieve successful seed germination, protocorm formation, and further seedling growth and development. This review provides both a basis for the selection of optimal conditions, and a platform for the discovery of better ones, that would allow the development of new protocols and the exploration of new hypotheses for germination and conservation of Dendrobium seeds and seedlings.

Transitions between self-compatibility and self-incompatibility and the evolution of reproductive isolation in the large and diverse tropical genus Dendrobium (Orchidaceae).

BACKGROUND AND AIMS: The evolution of interspecific reproductive barriers is crucial to understanding species evolution. This study examines the contribution of transitions between self-compatibility (SC) and self-incompatibility (SI) and genetic divergence in the evolution of reproductive barriers in Dendrobium, one of the largest orchid genera. Specifically, it investigates the evolution of pre- and postzygotic isolation and the effects of transitions between compatibility states on interspecific reproductive isolation within the genus. METHODS: The role of SC and SI changes in reproductive compatibility among species was examined using fruit set and seed viability data available in the literature from 86 species and ∼2500 hand pollinations. The evolution of SC and SI in Dendrobium species was investigated within a phylogenetic framework using internal transcribed spacer sequences available in GenBank. KEY RESULTS: Based on data from crossing experiments, estimations of genetic distance and the results of a literature survey, it was found that changes in SC and SI significantly influenced the compatibility between species in interspecific crosses. The number of fruits produced was significantly higher in crosses in which self-incompatible species acted as pollen donor for self-compatible species, following the SI × SC rule. Maximum likelihood and Bayesian tests did not reject transitions from SI to SC and from SC to SI across the Dendrobium phylogeny. In addition, postzygotic isolation (embryo mortality) was found to evolve gradually with genetic divergence, in agreement with previous results observed for other plant species, including orchids. CONCLUSIONS: Transitions between SC and SI and the gradual accumulation of genetic incompatibilities affecting postzygotic isolation are important mechanisms preventing gene flow among Dendrobium species, and may constitute important evolutionary processes contributing to the high levels of species diversity in this tropical orchid group.