As. Pac. J. Mol. Biol. & Biotech., July 2013 Vol. 3, 111-117
A Reliable Protocol for Micropropagation of Esmeralda clarkei Rchb.f. (Orchidaceae)
M.R. Paudel* and B. Pant
Plant Biotechnology and Biochemistry Laboratory, Central Department of Botany, Tribhuvan University, Kirtipur, Kathmandu, Nepal.
* Author for correspondence: M.R. Paudel
Plant Biotechnology and Biochemistry Laboratory, Central Department of Botany, Tribhuvan University, Kirtipur, Kathmandu, Nepal.
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Abstract.
A reliable protocol for micropropagation of Esmeralda clarkei Rchb.f., an epiphytic, ornamental vandaceous or¬chid was established. Mature seeds obtained from a green healthy capsule were inoculated in MS medium containing different concentrations and combinations of Benzylaminopurine (BAP) and 1-naphthaleneacetic acid (NAA) for established aseptic culture. The cultures were maintained at 25±2°C under a 16/8 hour light/dark photoperiod cycle provided by white fluorescent lamps (Philips, India). Protocorms were developed within 16 weeks; protocorms were differentiated into shoots after 25 weeks of seed culture in MS medium without added plant hormones. Among the different nutrient media tried in this study, BAP (0.5-2.0 mg/l) considerably induced multiple shoot growth (with 11 to 14 shoots growing 2.37-2.87 cm in length) from single protocorm explants and multiple shoots (9-11 shoots growing 3.75-3.87 cm in length) from single shoot section explants after 120 days of culture. More than 3 roots not less than 2 cm long were developed through in vitro rooting of shoots within 90 days of culture on MS medium containing NAA (0.5-1.0 mg/l) and various concentrations of plant hormone. In vitro grown plantlets were successfully acclimatized in hardening medium containing sand, soil and saw dust mixed in a 1:1:1 ratio.