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In Vitro Propagation of Dendrobium Lindeyii by Green Pod Culture

K. Kabir
Department of Horticulture, Sher-E-Bangla Agricultural University, Dhaka

K. A. Ara
Principal Scientific Officer
Floriculture Division, HRC, BARI, Gazipur

K., M. K. Samim
General Manager
Proshika Tissue Culture Lab., Koitta, Savar, Dhaka

M. R. Ali
Scientific Officer
Olericulture Division, HRC, BARI, Gazipur

Abstract/ Executive Summary:

Immature seeds obtained from green pod of Dendrobium lindeyii, an endangered epiphytic forest orchid having horticultural importance, can be germinated asymbiotically in vitro for rapid micropropagation. Murashige and Skoog medium containing 0.1 mg/l NAA and 15% coconut water was found most effective for high percentage (80-90%) seed germination, protocorm formation and seedling development. The advanced protocorm-like bodies when cultured on MS medium supplemented with 1.0 mg/l BAP was found to be optimum as indicated by highest shoot number (6.0) and shoot length (6.5 cm) at 90 DAI. The regenerated shoot were successfully rooted with maximum number (6.8) and longest rooting (6.5 cm) with 0.5 mg/l IBA supplemented with 0.5 g/l charcoal. The well rooted plantlets were transferred successfully to orchid house after hardening with 85% survival. This method can be exploited for their rapid micropropagation and conservation.

Keywords: Dendrobium lindeyii, green pod culture, in vitro propagation

Location: Department of Horticulture, Sher-E-Bangla Agricultural University, Dhaka

Start Date: 00-00-0000

End Date: 00-00-0000

Program Area: Seed Technology

Commodity: Orchid

Objectives:

To caring out for rapid mass propagation of D. lindeyii by asymbiotic in vitro green pod culture technique which can be exploited for their conservation.

Methodology:

Four to five months old green undehisced pods of D. lindeyii. was used as the source of explants for in vitro culture. The pods were cleaned thoroughly under tap water followed by washing with 5% Tween 20 for 10 min and finally with distilled water. They were surface sterilized with 0.1% mercuric chloride solution for 15 min and were subsequently rinsed 3 times in autoclaved distilled water. After surface sterilization, the capsules were taken in a sterile petridish containing filter paper to soak the surface water of the capsule. The capsules were cut longitudinally with a sharp sterilized surgical blade and with a long spatula powdery mass of yellowish seeds were scooped out and placed on basal MS media supplemented with 3% sucrose with several concentrations of naphthalene acetic acid (NAA) and 15-25% coconut water (CW) for seed germination. Difco-bacto agar (0.8%) was used as gelling agent and the pH of the media was adjusted to 5.6 using 0.1 N NaOH and 0.1 N HCl. About 15 ml of medium was dispensed in culture vessel and plugged before autoclaving at 1.05 kg cm^-2 pressure and at 121^oC for 20 min. The cultural vessels with inoculated seeds were kept in the culture room where 14 h of continuous light and 10 h of continuous dark period with 25+2⁰C were maintained. The cultures were monitored regularly. For each treatment 10 culture vials were used. The experimental design was completely randomized. The PLBs developed from the cultured immature embryos were maintained on the optimum germination medium for further differentiation. The PLBs with the first set of leaflets (advanced stage PLBs) were separated from the germination medium and cultured on regeneration media. For shoot multiplication, the MS medium was further supplemented with different concentrations of benzyl adenine purine (BAP) (0, 0.5, 1.0, 1.5 and 5.0 mg/l). For rooting, the resulted multiple shoot buds were separated and single shoots were placed on rooting medium containing ½ MS medium with 0.5 mg/l IBA supplemented with different concentrations of charcoal (0, 0.2, 0.5, 1.0 and 2.0 mg/l). Data were recorded at 30, 60 and 90 DAI for both shoot proliferation and rooting. The well rooted plantlets were maintained for 2-3 passages on the regeneration medium before they were transferred to hardening condition. About 5-7 cm long rooted plantlets with 3-4 leaves and 4 or 5 roots with distinct pseudobulb from the regeneration medium were used for the hardening. The plantlets were maintained in culture vials containing 1/10^th MS basal medium with sucrose (1%). Sterilized cocodust and charcoal (1:1 ratio) were also incorporated as substratum in the culture vial and maintained for 5-6 weeks before transferring to potting mix. The potted plants were exposed to normal day light for about 1 h in a day for initial one weekend subsequently increased the exposure period by 2 h from the second week and after one month the plantlets were left in the normal full day light condition. Finally, the potted plants were gradually shifted to the orchid net house.

Source of Information: Bangladesh J. Seed Sci. & Tech. 16 (1 &2): 21-27 (2012),ISSN 1029 - 8800

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

The fully developed plantlets were then transferred to orchid house for hardening (Plate 1e). and gradually the plantlets grew well (Plate 1f). About 85% survival rate was registered after 3 months of potting. The result of the present investigation indicated that, green pod culture techniques could be formulated for germination, mass scale micro propagation and production of plantlets of Dendrobium lindeyii within a short time. This strategy offers an opportunity for using the techniques in commercial nurseries.

Knowledge Management: Journal