Optimization of Embryogenic Callus Induction and Plant Regeneration in Orchid Coelogyne cristata

An efficient protocol was established for high frequency somatic embryogenesis through a callus culture of Coelogyne cristata. The best frequency of callusing was obtained from a PLB segment (3-5 mm) cultured on MS medium supplemented with coconut water (CW) and a combination of both 3 $mg{\cdot}L^{-1}$ of 2,4-D and BA. When the calli were sub-cultured on the MS medium without any PGRs, the average number of somatic embryos were higher than those with PGRs treatment. NAA is the most critical factor among PGRs, which dramatically hindered for the formation of a somatic embryo. The efficacy of the addition of coconut powder (CP) for somatic embryogenesis was almost the same in all treatments. However, the number of somatic embryos formed distinctly depended on age of the callus. The somatic embryos converted into healthy plants with well-developed shoots on the same medium. Plantlets showed the best responses of root and shoot growth when transferred to $\frac{1}{2}$ MS medium containing 1.5 $g{\cdot}L^{-1}$ of activated charcoal. All plants with above 3.0-cm-high were successfully acclimatized in the greenhouse.