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Assessment of Somatic Embryogenesis and Plant Regeneration Potentiality from Coleoptile and Root Tissues of Jhum Rice (oryza Sativa L.)

Pervin Akter
Department of Botany, University of Chittagong, Chittagong- 4331, Bangladesh.

M. AL-Forkan
Department of Botany, University of Chittagong, Chittagong- 4331, Bangladesh.

Abstract/ Executive Summary:

In this study, attempt was made to establish a reproducible protocol via tissue culture from root and coleoptiles in three Jhum rice cultivars (cvs.) viz Hamaran, Sori, Kulbadam. Spontaneous callus induction and plant regeneration as well as variation among the cultivars were observed in different combinations on MS based media. Embryogenic calli and non embryogenic calli were found on MS based media supplemented with 2.5 mgl^-1 2, 4-D; 1% Casein hydrolysate (CH) and 0.5 mgl^-1 Kn respectively. Plantlet regeneration occurred on MS basal medium using 2.0 mgl^-1 BAP and 0.5 mgl^-1 Kn. In point of coleoptiles, cv. Hamaran displayed a good number of embryogenic calli and shoot production. Most effective response amongst the cultivars was found in coleoptiles than root while explants of root failed to reproduce plant lets through tissue culture. Finally, the plant lets were transferred to the natural condition in pot for acclimatization.

Keywords: Jhum rice, Coleoptiles, Roots, Embryogenesis, Plant regeneration

Location: Department of Botany, University of Chittagong

Start Date:

End Date:

Program Area: Crop-Soil-Water Management

Commodity: Rice

Objectives:

To determine the response of callus production and regeneration efficiency of root and coleoptiles from some hilly cultivated jhum rice (Oryza sativa L.).

Methodology:

Seeds of three jhum rice cultivars (Oryza sativa L) viz. Hamaran, Sori and Kulbadam were used in this study. Seeds were carefully dehisced manually, dipped in 70% ethanol for 2-3 min, and then immersed in 0.2% HgCl₂ solution for 20 min with regular agitation. Then the seeds were rinsed for 4-5 times with doubled distilled water to remove HgCl₂ from surface. After that, seeds were placed on 70 mm sterile filter paper for about 10 min for removing excess water. Then the sterile seeds were kept on MS (Murashige and Skoog, 1962) basal media devoid of growth hormones in the dark at 26°C. Only eight-day-old root and 5 day old coleoptiles were taken aseptically for culturing. The base and tips were removed on both the explants. Five to seven pieces of root coleoptiles were cultured separately in each Petri dish on callus induction media kept at dark chamber for 14 days at 26 ± 2°C. The subcultures were maintained after 20-25 days interval with transfer of only embryogenic calli. Cultures showing infectious, slow growth, more rhizogenic, watery calli were removed immediately in each subculture. The MS based media with different combinations such as 2.5 mgl^-1 2, 4-D; 2.5 mgl^-1 2, 4-D + 1% CH ; 2.5 mgl^-1 2, 4-D + 0.5 mgl^-1 kn enriched with 30 gl^-1 (w/v) sucrose and gelled with 0.8% (w/v) agar to solidify the media were used for initiation of callus. The MS based plant regeneration media such as 2.0 mgl^-1 BAP + 0.5 mgl^-1 Kn with 3% (w/v) sucrose was used where a part of the media were semi solidified with 1% agar (w/v) for initial culture of embryogenic calli (containing 4-6 segments of embryogenic calli) were kept in the dark at 26 ± 2°C for 18 days prior to transfer in the light condition. Then calli were cultured (after 10 d) on the rest of other portion of plant regeneration media in which 0.8% (w/v) agar was used to solidify the media. After the next 10 d of cultured calli turned greenish and then gradually it developed plant lets with shoot and root within one and half months. Each 2-3 cm in height of plant lets were taken from individual derived calli and multiplied by transferring (20-25 d) to 175 ml capacity of reagent conical flask containing 40 ml of MS based rooting medium supplemented with 1.5 mgl^-1 NAA and 3% (w/v) sucrose semi solidified with 0.8% (w/ v) agar. The pH was always adjusted to 5.8 with 0.1N HCL or 0.1N NaOH before adding the agar with media. The media after preparation and the culture vessels were autoclaved at 1.9 kg/cm 2 of pressure at a temperature of 121°C for 20 min for sterilization. Normal and well developed plant lets with vigorous root of plant lets were preferred to plant in the natural pot culture in pot. For acclimatization, in vitro derived plant lets were kept under shade or covered with porous polythene bag is gradually exposed to sunlight. In this study, germination capability of seeds, frequency of callus ability (embryogenic and non-embryogenic), the regeneration frequencies, callus size, callus growth were recorded accordingly during maintenance in each subculture.

Source of Information: Indian J. Agric. Res., 44 (2) : 88 - 95, 2010

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

The investigation showed that the production of embryogenic callus in both roots and coleoptiles were observed whereas the maximum number of embryogenic callus produced by coleoptiles. Moreover, the type of callus development was widely varied among the variety tested in media containing different concentrations. It may be assumed that callogenesis and regeneration potential depend upon genetic constitutes as well as environmental reaction and nutrition media. Although root could produce embryogenic callus but failed to reproduce plantlet. On the other hand embryoids of coleoptiles have shown its totipotency which may be a base line of tissue culture for genetic manipulation for the improvement of rice.

Knowledge Management: Journal