Development and characterization of flower and capsule in Borneo jewel orchid Macodes limii J.J. Wood and A.L. Lamb (Orchidaceae: Asparagales)

Macodes limii J.J. Wood and A.L. Lamb is an endemic jewel orchid of Sabah in Borneo. The species has unique characteristic with high potential to be commercialized as ornamental plant. However, it was understudied because of its scarcity in nature, which can only be found at certain area in Sabah with ultramafic soil. To understand the reproduction biology of this species, the present study was conducted to document the developmental stages of flower and capsule to determine the right harvesting time for further use in seed germination study. Macodes limii was collected from its natural habitat and cultivated ex situ throughout this study. Flower development was recorded starting from inflorescence initiation until full bloom, which took over 54 days to complete. Flower was artificially pollinated, and capsule was produced within 22 days, and harvesting was performed prior to dehiscence. Seeds were aseptically cultured on half-strength Murashige and Skoog medium þ 3% (w/v) sucrose, and 29% of seeds were successfully germinated after 24 weeks under continuous dark condition. The current findings are beneficial as a reference to support the conservation efforts of this endemic specie

Effect of basal media and carbon sources on callus culture maintenance of Vanda dearei

Vanda dearei is an endemic orchid of Borneo and has been listed as an endangered orchid in Appendix II of the Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES). Vanda dearei has beautiful pale yellow-flowers, large petals and strongly scented. Therefore, in vitro micropropagation has been applied in order to develop a novel micropropagation method to mass produce this species. Through callus culture techniques, orchids with limited resources can be mass propagated in a shorter period. However, callus culture in orchid is hardly maintain due somatic embryogenesis properties and easily regenerated to plantlets. Thus, this study aims to develop an efficient protocol for callus cultures of V. dearei by manipulating basal media strengths and carbon sources. Callus induced from the leaf segments of V. dearei were used as explants and were cultured on KC, Mitra, MS and VW basal medium at different nutrient strengths (1/4, 1/2, 1 and 2x) added with 1.0:0.1 mg/l TDZ:NAA and 1 to 4% (w/v) of sucrose, glucose or fructose, respectively. All cultures were incubated in the dark with temperature of 25±2°C. Results showed that callus growth has improved with decreased nutrients strength of basal media. Quarter strength of Mitra medium promotes the best condition for callus maintenance to approximately 8.00±17.89% at 8 weeks of culture. This is followed by the ½ strength of MS and ¼ strength of VW with 8.00±10.95% and 5.00±10.00%, respectively. Callus grown on the other basal strengths are mainly differentiated and developed into protocorm like bodies (PLBs), especially at double strengths (100±0% explants turn into PLBs). In addition, low percentage of necrosis (less than 28%) was also observed on Mitra basal medium compared to the other media (more than 36%). Sucrose has been identified as the best carbon source to support callus growth followed by glucose and fructose. Addition of 1% (w/v) sucrose increased callus maintenance up to 32±17.9%, promote cell differentiation and increased average size of callus (1.52±0.63 callus score). This treatment also support the longest retention time of explant maintained in callus for 5 weeks and has the lowest percentage of callus necrosis (20±24.5%)

Effects of Organic Additives and Plant Growth Regulators on Protocorm Development of Dendrobium lowii

A simple and efficient growth protocol was develop forDendrobium lowii,an endangered and Borneo’s endemic epiphyte orchid, using four month old protocorms as explant sources produced by asymbiotic seeds germination. Protocorms of Dendrobium lowii were cultured on Knudson C (KC) media supplemented with organic additives (coconut water, tomato juice and banana pulp) or plant growth regulators (NAA, Zeatin and BAP) at different concentrations and observed for protocorm development. Among all organic additives tested, medium containing banana pulp at 25g/L induced the highest growth index value of 593.3 after 240 days of culture. This treatment also promoted 100% production of shoot and 93.3% of root formation compared to other treatments. Addition of 2g/L peptone or 15% (v/v) coconut water had significantly induced 16.7% protocorms proliferation. The supplementation of 6 μM NAA promotes similar responses for growth index of 563.3. The treatment induced up to 86.7% and 83.3% of protocorms forming shoots and roots, respectively. The study also revealed that the addition of 2 or 4 μM of NAA and 4 or 6 μM BAP is suitable for shoot induction, however with poor rooting formation. This finding is important for conservation and horticultural manipulation of the species