Agricultural Research Management Information System

Home
Research Summary
Search
About Us
- ARMIS
- Brochure
Contact Us
Report
User Request
Data Input
Help
- Operation Manual
  - PDF
  - Video
- Program Area & Commodity
We have reached 37600 number of research entries at this moment.

- Logout

Research Detail

Home
Research
Detail

High Frequency Regeneration of Phalaenopsis Amabilis (l.) Bl. Cv. Lovely Through in Vitro Culture

Pinaki Sinha
Biological Research Division, BCSIR Laboratories, Dhaka

Miskat Ara Akhter Jahan
Biological Research Division, BCSIR Laboratories, Dhaka

John Liton Munshi
Biological Research Division, BCSIR Laboratories, Dhaka

Rahima Khatun
Biological Research Division, BCSIR Laboratories, Dhaka

Abstract/ Executive Summary:

Young leaf explants from mature plant of Phalaenopsis amabilis cv. Lovely were cultured on half strength of MS supplemented with BA (2.0 mg/l), NAA (0.5 mg/l), 2% (w/v) sucrose, 10% (v/v) coconut water, 2 g/l peptone and 1 g/l activated charcoal. Each section of explant produced ten protocorm‐like bodies (PLBs) after 12 weeks of culture. The PLBs were subcultured on the same nutrient medium but without phytohormone and addition of 150 mg/l L‐glutamine, where PLBs were found to be enlarged with leafy shoots and new PLBs were induced from the base of the old ones. Plantlet development from leafy shoots was achieved on half strength MS supplemented with 2 g/l peptone, 2% (w/v) sucrose, 10% (v/v) CW and 1 g/l activated charcoal, where 100% explants were developed into plantlets with roots within eight weeks. The addition of 2.5 g/l banana powder enhanced the number and length of roots. Within the first 32 weeks after initiation of culture about 1200 plantlets as well as a huge amount of PLBs were achieved from a single explant section. The plantlets were acclimated in natural environment.

Keywords: Orchid, Phalaenopsis amabilis, Protocorm, Clonal propagation

Location:

Start Date:

End Date:

Program Area: Knowledge Management

Commodity: Orchid

Objectives:

This paper reports an efficient and quick method for continuous high frequency clonal propagation of purple coloured P. amabilis cv. ʹLovelyʹ through in vitro culture of young leaf sections of mature plants.

Methodology:

Young emerging leaves of the purple cultivar of Phalaenopsis amabilis (L.) Bl. cv. ʹLovelyʹ were collected from the nursery and used as explants. The young leaves were washed under running tap water followed by detergent, Tween 80 (5% v/v) for 5 min. After a thorough wash with double‐distilled water, surface sterilization was done with 0.1% (w/v) HgCl₂ solution for 8‐10 min. The explants were again washed thoroughly with sterile double‐distilled water. The surface sterilized leaves were then prepared for inoculation by cutting into pieces. For direct induction of PLBs, the explants were cultured on half strength of MS supplemented with BA (0.5 ‐ 2.5 mg/l) and Kn (0.5 ‐ 2.5 mg/l) individually and in combination with NAA (0.2 ‐ 0.5 mg/l) along with 2% sucrose, 10% (v/v) coconut water (CW), 2 g/l peptone and 1 g/l activated charcoal. The medium was gelled with 2.2 g/l gelrite (Duchefa, The Netherlands). The pH of the media was adjusted to 5.6 before autoclaving at a pressure of 105 kPa for 20 min at 121°C. All cultures were incubated at 24 ± 1°C, under cool white fluorescent light of 30 μmol m2/s for 16 hr per day. For proliferation of PLBs and formation of shoots, each clump of PLBs developed from initial explants was cut into four pieces and subcultured on two different media, MS and half strength of MS supplemented with 2% sucrose, 2 g/l peptone, 1 g/l activated charcoal and with or without coconut water (10% CW) and L‐glutamin (50 ‐ 300 mg/l). In all experiments explants were cultured in 250 ml conical flasks or disposed off jam bottles (86 × 120 mm) containing 40 ml medium. In PLB proliferation medium, old PLBs were developed into leafy structures from the base of which new PLBs emerged. These leafy structures or shoots were subcultured on half strength of MS with 2% sucrose, 2 g/l peptone, 10% CW and 1 g/l activated charcoal for plantlet formation. Banana powder was also used to study its efficacy in root differentiation and growth. For acclimatization the rooted plantlets (between 50 and 70 mm in height) were taken out from the media and washed with tap water to remove the gel adhered to the roots. They were then implanted in a plastic basket containing coconut husk. To maintain high humidity the plantlets were misted twice a day. After acclimatization the plantlets were transplanted to 14 cm³ plastic pots perforated at the bottom and containing coconut husk (20 mm³) and charcoal (10 mm³) (2 : 1) and maintained under shade at 30/25°C (day/night). Plants were watered every alternate day and fertilized with 6.5N‐4.5P‐19K solution at ten days intervals. Experiments were performed in a randomized design and all experiments were repeated three times. In in vitro culture each treatment had 15 replicates. The morphogenetic response of explants for PLB induction was evaluated after 12 weeks of culture. For PLB proliferation and plantlet regeneration, results were evaluated within eight weeks of culture. Data were statistically analyzed and means were compared using DMRT (Duncan 1955).

Source of Information: Plant Tissue Cult. & Biotech. 20(2): 185-193, 2010 (December)

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

In conclusion, an efficient protocol for continuous high frequency regeneration of pink cultivar of P. amabilis cv. ʹLovelyʹ in a simple culture medium and short culture period was established. The authors have optimized the high frequency PLB proliferation medium (half strength of MS in combination with 2% (w/v) sucrose + 2 g/l peptone + 1 g/l activated charcoal + 10% (v/v) CW and 150 mg/l L‐glutamine) and method (culture of PLB clump, instead of PLB section) and conversion of leafy shoots into plantlets in simple half MS. Likewise, through repeated subculture of PLB clumps, removing of leafy shoots and their subsequent subculture to plantlet regeneration medium and harvesting of regenerated plantlets, continuous high frequency production of plants could be maintained.

Knowledge Management: Journal