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In Vitro Regeneration of Arabioopsis Thaliana

M. K. Amin
Deparrnent of Biotechnology, Bangladesh Agricultural University, Mymensingh

M. A. Islam
Department of Genetics and Plant Breeding, Bangladesh Agricultural University, Mymensingh

G. H. M. Sagor
Department of Genetics and Plant Breeding, Bangladesh Agricultural University, Mymensingh

L. Hassan
Department of Genetics and Plant Breeding, Bangladesh Agricultural University, Mymensingh

Abstract/ Executive Summary:

The experiment was conducted, to develop a suitable and reproducible protocol and for efficient in vitro regeneration of plantlets from the leaf and root explants of Arabidopsis thaliana. Investigation was also done to determine the regeneration potential of the explants used in this experiment. After germination in MS medium leaves from 15 days old plants and roots from 21 days old plants were used as explants for callus induction in basic medium with different concentrations of 2,4-D and constant concentration of IAA and Kinetin. The highest callus induction (92%) was observed from root in the media supplemented with 0.5 mg/L 2,4-D, O.3mg/L Kinetin and 5 mg/L IAA followed by 1 mg/L 2,4-D, 0.3 mg/L kinetin, and 5 mg/L IAA. Shoot regeneration was highest (31.25%) from the calli derived from root in the basic media supplemented with 0.15 mg/L IAA, 2 mg/L BAP and 5 mg/L 2iP. Highest root initiation (80%) was observed in the shoot derived from root explants in half MS medium supplemented with 0.5 mg/L NAA. The in vitro regenerated plantlets from root explants were successfully established in field condition.

Keywords: Arabidopsis thaliana, Regeneration, 2,4-D, BAP,

Location: Tissue Culture laboratory of the Department of Genetics and Plant Breeding, BAU, Mymensingh

Start Date: 01-06-2005

End Date: 01-12-2005

Program Area: Knowledge Management

Commodity:

Objectives:

It is very important plant from biotechnological point of view, but still it is not habituated in Bangladesh- For this reason, it is essential to introduce this plant in our country and find out the suitable protocol of in vitro regeneration. Subsequently, this will help to grow this plant species in existing field condition and it would be possible to do the research on molecular and genetic level. Keeping the facts in mind, this piece of work was undertaken.

Methodology:

Seeds of Arabidopsis thaliana were obtained by the courtesy of Centre for Cellular and Molecular Biology (CCMB), Hydrabad, India and the experiment was conducted during the period from June-December, 2005 at the Tissue Culture laboratory of the Department of Genetics and Plant Breeding in Bangladesh Agricultural University, Mymensingh. The seeds were sterilized by 1 % sodium hypochlorite solution followed by through washing with sterile distilled water. Sterilized seeds were transferred into plates containing germination media placing maximum 5 seeds in each plate. The plates were incubated in dark for 5 to 7 days until seeds were germinated. On germination the plates were transferred into light. Leaves of 15 days old and roots of 21 days old seedling of in vitro grown plants were used as explants for callus induction and cultured on basic medium supplemented with different concentrations of 2,4-D (0, 0.5, 1.0 and 2.0 mg/L) and constant concentration of IAA(5.0 mg/L) and Kinetin(0.3 mg/L). Callus was formed within 3-4 weeks and at this stage the callus was sub-cultured for shoot initiation on basic medium supplemented with different concentrations of BAP (2.0, 3.0, and 3.5 mg/L) and a constant concentration of IAA (0.15 mg/L) and 2iP (5.0 mg/L). Regenerated plantlets were sub-cultured in half MS (Murashige and Skoog, 1962) medium with different concentrations of NAA (0, 0.5 and 1.0 mg/L) for root formation and finally the plants were established in existing field condition.

Source of Information: Bangladesh j. crop sci. 2006, 17 (1): 181-188

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

Different concentrations of BAP showed Significant variations for number of callus showing shoot/vial and days to shoot initiation, indicating important differences among the different level of BAP on these characters. BAP at 2.0 mg/L concentration was found to be the best, for number of callus showing shoot/vial but BAP at 3.0 mg/L was found best for days to shoot initiation. Days to shoot initiation was minimum (21 days) at 3.0 mg/L BAP and maximum (32.4 days) at 2.0 mg/L BAP. Effects of explants on shoot induction, mean values of two explants were found statistically significant for all the characters of shoot regeneration like number of callus showing shoot/vial and days to shoot initiation. Number of callus showing shoot/vial was the highest in root explants (0.833). It was observed that among the explants tested, root explants showed minimum number of days to shoot initiation (25.714). It is evident from the studies that callus derived from root explants showed the better performance for different characters. Effects of hormone x explants on shoot induction, the differences among the interactions for these characters were statistically significant. Number of callus showing shoot/vial was the highest on T₁ and root explants (1.25), whereas, no shoot was found in T ₃ and leaf explants. Days to shoot initiation was minimum (20.750) in T₂ and root explants and maximum (32.500) in T₃ and leaf explants. Callus initiation of Arabidopsis from root explants (A) and leaf explants (B) on basic media + 1.0 mg/L 2,4-D + 0.3 mg/L kinetin+ 5 mg/L IAA. Shoot initiation of Arabidopsis from leaf explants (C) and root explants (D) on basic media + 0.15 mg/L IAA + 5 mg/L 2iP + 2 mg/L BAP. Initiation of root from regenerated shoot of Arabidopsis on 1/2 MS + 0.5 mg/L NAA (E). Established plantlet of Arabidopsis in pot (F).

Knowledge Management: Journal