2016 Volume 33 Issue 3 Pages 153-160
The paper describes an efficient and reproducible protocol for in vitro propagation of Herminium lanceum, a medicinal orchid, natural polulations of which are adversely affected because of its indiscriminate collections and other anthropogenic activities. The germination of seeds at different stages of embryo development was compared on four media: Mitra’s (M), M +0.1% peptone (M+P), 1/2 Murashige and Skoog’s+0.1% peptone (MS+P) and Knudson C+0.1% peptone (KC+P). The seeds that contained multicellular oval embryos and cultured on M+P exhibited the highest percentage (82.94%) of germination. Protocorm-like bodies (PLBs), developed from the seeds, could be multiplied by repeated sub-cultures on M+P (hereafter referred to as basal medium, BM). PLBs sub-cultured on BM, supplemented with different concentrations of 6-benzylaminopurine (BAP), kinetin (KN) or 1-phenyl-3-(1, 2, 3-thiadiazol-5-yl)-urea (TDZ) differentiated shoots. The maximum number of shoots per culture developed on BM+4 µM TDZ. Shoot elongation was the best on BM+0.1% AC+1 µM GA3. Among various media tested for rooting of the shoots, the best rhizogenic response was obtained on BM+4 µM TDZ+0.1 µM IBA. About 82% of in vitro regenerated plantlets survived on transfer to the potting mixture of sand : vermiculite (1 : 1). High performance liquid chromatographic (HPLC) analyses revealed the presence of medicinally important phenolic acids in leaves and tubers of both in vitro and in vivo plants. The developed protocol could be used for the large-scale production of propagules required for the cultivation of this plant as a regular source of herbal, and also for eco-restoration of its native habitats.
