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In Vitro Plant Regeneration from Cotyledon and Internodes Derived Callus in Watermelon (citrulus Lanatus Thumb.)

M. M. Khatun
Department of Genetic Engineering and Biotechnology, University of Rajshahi, Rajshahi-6205, Bangladesh

M. S. Hossain
Department of Genetic Engineering and Biotechnology, University of Rajshahi, Rajshahi-6205, Bangladesh

M. Khalekuzzaman
Department of Genetic Engineering and Biotechnology, University of Rajshahi, Rajshahi-6205, Bangladesh

A. Rownaq
Department of Genetic Engineering and Biotechnology, University of Rajshahi, Rajshahi-6205, Bangladesh

M. Rahman
Department of Genetic Engineering and Biotechnology, University of Rajshahi, Rajshahi-6205, Bangladesh

Abstract/ Executive Summary:

A study was carried out to observe the callus induction and subsequent regeneration potentiality of watermelon (Citrullus lanatus Thumb.) from cotyledon and internodes of this plant. Greenish compact callus was achieved from cotyledon on MS medium supplemented with 1 mg/l 2, 4-D within one week of inoculation. The callus produced large number of shoots when cultured on MS medium with1.0 mg/l BAP+0.2 mg/l NAA. Rooting rate was 100% when shoots from second subculture were cultured in medium with 1.0 mg/1 IBA and the shoots were more prone to rooting by increasing subcultures. In vitro grown plantlets with well root system were successfully established in natural condition through successive phases of acclimatization. The regenerated plantlets were healthy, uniform and identical to donor plants and survival percentage was 80%.

Keywords: Callus induction, Plant regeneration, Cotyledon and internodes, Citrullus lanatus

Location: Professor Ali. Md. Eunus Laboratory, Dept. of Genetic Engineering & Biotechnology, University of Rajshahi, Rajshahi-6205

Start Date: 00-10-2007

End Date: 00-02-2008

Program Area: Crop-Soil-Water Management

Commodity: Water melon

Objectives:

To observe the callus induction and subsequent regeneration potentiality of watermelon (Citrullus lanatus Thumb.) from cotyledon and internodes of this plant.

Methodology:

The present experiment was carried out at the Professor Ali. Md. Eunus Laboratory, Dept. of Genetic Engineering & Biotechnology, University of Rajshahi, Rajshahi-6205, Bangladesh during October to February. The seeds of the crop (Citrullus lanatus Thumb.) was collected from seed market, Siddique Bazar, Dhaka. These seeds were packed by TAKII and CO., LTD. Kyoto. Japan. The seeds were taken in a conical flask and thoroughly washed under running tap water for 30 min. to reduce the level of surface microorganisms and to loose the seed coat. Then the seeds were taken in reagent bottle containing distilled water with few drops of tween-80 (wetting agent) and 2-3 drops of savlon for about 10-12 minutes rinsed with constant shaking. This was followed by a second washing with distilled water to remove all traces of above chemicals. Surface sterilization was carried out by 0.1% HgCl₂ with gentle shaking for solution for 5 minutes. Then the sterilized seeds were washed 5-7 times with sterile distilled water immediately to remove all traces of HgCl₂. This procedure was carried out in aseptic condition of laminar airflow cabinet. The sterilized seeds were then inoculated into MS medium. After one week, in vitro grown seedlings were the ready sources of different kinds of explants like, shoot tips, nodal segments cotyledons, and internodes which were free from contamination. In the present study, cotyledon and internodal explants were collected from aseptically germinated seedlings for callus induction and plant regeneration. The explants were cultured on MS medium with 3% (W/V) sucrose which was solidified with 0.7% (W/V) agar. The pH of the media was adjusted to 5.7 prior to autoclaving at 121^oC for 20 minutes. The cultures were incubated in a culture room at 25±2^oC with a photoperiod of 16 hour at 3000 lux light intensity provided by cool white fluorescent tubes. For each treatment, 10 replications were used and all experiments were repeats thrice. Visual observation of culture was made in every week. Data on callus, shoot and root induction were recorded after 30, 35 and 25 days of inoculation respectively. The medium supplemented with different concentrations of NAA (Naphthalene acetic acid) and 2, 4-D (2,4-Dichlorophenoxy acetic acid), in order to obtain the most suitable culture media for induction of high frequency of callus. Once the callus developed, they were further cultured for regeneration and elongation of shoots in the medium having different concentrations of auxins and cytokinins. Elongated shoots were rooted on MS medium supplemented with different concentrations of auxins (Naphthalene acetic acid and Indole-3 butyric acid) singly. After proper rooting, the plantlets were gradually exposed to normal conditions for acclimatization and transferred to natural condition where the plantlets were grown successfully.

Source of Information: Int. J. Sustain. Crop Prod. 5(4):25-29, 2010

Article Link (Online Journal):

Funding Source:

1. Government

Budget:

Total Budget (Tk.) :

Achievement/Findings:

In the present study, it was observed that 2, 4-D was found as the best auxin for callus induction in Citrulus lanatus Thumb. Again, the cotyledon induced callus performed better than the internodal callus for shoot multiplication which justified its further use in root induction. These results indicated that cotyledon might be used for suitable source of explants for the improvement of this crop through biotechnological approaches.

Knowledge Management: Journal