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In Vitro Micropropagation of Rauvolfia Serpentina (l.) Benth Through Induction of Direct and Indirect Organogenesis

S. K. BHADRA
Department of Botany, University of Chittagong, Chittagong - 4331, Bangladesh

P. SINGH
Department of Botany, University of Chittagong, Chittagong - 4331, Bangladesh

T. K. BHOWMIK
Department of Botany, University of Chittagong, Chittagong - 4331, Bangladesh

Abstract/ Executive Summary:

Leaf and nodal segments of two months old field grown seedlings of Rauvolfia serpentina (L.) Benth were aseptically cultured on agar solidified MS medium supplemented with various combinations and concentrations of auxins (NAA, IAA, 2,4-D and picloram) and cytokinins (BAP and Kn). The nodal segments produced highest number of multiple shoot buds (5.85/explant) on MS medium supplemented with 2.0 mgl^-1 BAP + 0.2 mgl^-1 NAA or 2.0 mgl^-1 BAP + 0.1 mgl^-1 IAA. Whereas nodal segment produced callus tissue of different nature on MS medium supplemented with 1.5 mgl^-1 BAP + 0.5 mgl^-1 IAA+ 1.5 mgl^-1 2,4-D; 3.0 mgl^-1BAP + 1.0 mgl^-1 NAA + 1.5 mgl^-1 Kn and 0.1 mgl^-1 Pic + 1.0 mgl^-1 Kn. The callus tissue of light green and nodular nature on further subculture in a wide range of plant growth regulators (PGRs) supplemented media, differentiated into multiple shoot buds that underwent rapid elongation on 2.0 mg/l BAP and 0.2 mg/l NAA supplemented media. The elongated shoot buds on further subculture in rooting media produced strong and stout roots. Half strength MS with 1.5% (w/v) sucrose was most effective for enhancing rooting. Finally those plantlets were acclimatized in field. Thus a protocol was established for rapid micropropagation of this medicinal plant through induction of direct and indirect organogenesis from nodal explant.

Keywords: Micropropagation, Rauvolfia Serpentina, Organogenesis

Location: Botany Deparment of Chittagong University

Start Date:

End Date:

Program Area: Development of Host and Medicinal Plants

Commodity: Plant

Objectives:

1. The research study was undertaken with a view to develop a reliable, reproducible and efficient protocol for in vitro propagation of the medicinal plant species.

Methodology:

Two months old seedlings were collected from the nursery of BCSIR, Chittagong, and were established in garden pots of Botany Deparment of Chittagong University. Leaf and nodal segments of approximate 0.5 - 1.0 cm size were used as explant. Surface sterilization of the leaves and nodes was done by dipping in 70% ethanol for 30 sec. and submerging them in 0.2 % (w/v) HgCl₂ solution for 10 min. The segments were then washed three times with sterile distilled water and finally cultured in test tube (2.5 × 15 cm) and conical flask (100 ml). MS basal medium supplemented with various combinations of plant growth regulators (PGRs), viz., auxins (NAA, IAA, 2,4-D and Picloram) and cytokinins (BAP and Kn) at different concentrations were used for culture of the explants. All the media were made 0.8% (w/v) agar solidified and pH of the media was adjusted to 5.8 prior to autoclaving for 20 min. at 121⁰C under 1.9 kg/cm² pressure. Culture vessels with inoculated explants were maintained in a culture room under a regular cycle of 14h light and 10h dark at 25 ± 2⁰C. The nodal segments underwent direct or indirect organogenesis in some of the media combinations giving rise to multiple shoot buds. The multiple shoot buds were further grown on elongation media and finally the elongated shoot buds were individually rooted on rooting media. The complete plantlets were finally transferred and established in outside natural environment through successive phases of acclimatization.

Source of Information: The Chittagong Univ. J. B. Sci.,Vol. 3(1 &2):pp.01-09 , 2008.

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

For induction of strong and stout root system the elongated shoots were individually grown on four different rooting media, viz., (i) half strength MS + 1.5% (w/v) sucrose (ii) half strength MS + 1.5% (w/v) sucrose + 0.5 mg/l IAA + 0.5 mg/l IBA (iii) half strength MS + 1.5% (w/v) sucrose + 0.5 mg/l NAA and (iv) full strength MS + 3% (w/v) sucrose + 0.5 mg/l IAA. Half strength MS with 1.5% (w/v) sucrose was found most effective for rooting. It proved, the auxin free medium was best for rooting. Well developed plantlets were then transferred to outside under natural environment through sequential phases of acclimatization. The protocol thus developed for mass production of seedlings of R. serpentina can be used reliably for conservation of this important medicinal plant species of Bangladesh.

Knowledge Management: Journal