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Plant Regeneration Through in Vitro Cormel Formation from Callus Culture of Gladiolus Primulinus Baker

Pinaki Sinha
Department of Botany, Jahangirnagar University, Savar, Dhaka, Bangladesh

Shyamal K. Roy
Department of Botany, Jahangirnagar University, Savar, Dhaka, Bangladesh

Abstract/ Executive Summary:

Callus capable of plant regeneration was initiated at a higher frequency from slices of cormel sprouts of Gladiolus primulinus cv. Golden Wave, cultured in MS with 4.0 mg/l NAA, 2.0 mg/l BAP or 1.0 mg/l 2,4-D. Callus was maintained in callus initiation medium. Shoots regenerated from the callus in MS with 0.5 - 2.0 mg/l Kn or/and 0.5 - 2.0 mg/l BAP. Highest number of shoots regenerated in MS with 2.0 mg/l BAP from one-month-old callus which was initiated in the medium with NAA. Shoot regeneration capacity of callus decreased with the age and after fifth month no shoot was found to be regenerated. The shoots produced roots in half strength of MS with 2.0 mg/l IBA and 6 % sucrose. Rooted shoots produced cormels at the base of shoots in the same nutrient medium. About 80 - 90 % cormels germinated when sown in the field and 20 % of them blossomed in the first year.

Keywords: Gladiolus, Ornamental, Regeneration, Cormel formation

Location: The tissue culture laboratory, Jahangirnagar University, Dhaka, Bangladesh

Start Date:

End Date:

Program Area: Variety and Species

Commodity: Gladiolus

Objectives:

To determine the proper concentrations of growth regulators for callus initiation and regeneration of a locally cultivated Gladiolus cv. Golden Wave, taking young cormel sprout as explant.

Methodology:

For sprouting the cormels were aseptically cultured in half strength of MS with 1,0 mg/l BAP. Before culture the cormels were surface sterilized with continuous stream of running tap water for 30 min, washed with liquid detergent for 15 min and then with a solution of the antiseptic savlon 5 % (v/v) for 10 min. The cormels were then washed repeatedly with distilled water and finally treated with HgCl₂ (0.2 %) for 10 min in a laminar air flow cabinet and washed for two times with autoclaved double distilled water to remove any trace of HgCl₂. MS containing standard salts and vitamins, 3 % (w/v) sucrose and 0.7 % (w/v) agar was used. The pH of the medium was adjusted to 5.8 before adding agar. The medium was autoclaved at 1.1 kg/cm2 for 20 min at 120_C. Cultures were incubated at 10,000 lux with cool white fluorescent light and maintained at 24 ± 2_C. Callus was initiated from seven-day-old sprouts of cormels cultured in vitro. Each sprout was sliced into 2 mm thick transverse sections and they were placed with a cut surface in contact with culture medium. The medium for callus initiation was BMS B basal salts, 3 % (w/v) sucrose, 0.7 % agar (BDH), and the followings in mg/l : glycine, 2.0; thiamine, 1.0; pyridoxine, 0.5; nicotinic acid, 0.5; mesoinositol, 100.00 and various hormones, like 2,4-D, BAP and NAA, individually at different concentrations. One gram of each fresh weight samples of callus were placed on shoot induction medium. Each treatment consisted of five callus samples. Medium for shoot induction was MS supplemented with Kn and/or BAP in different concentrations. When the regenerated shoots were 2 - 3 cm tall, they were separated from the base and implanted in half strength of MS with IBA (0.5 - 2.5 mg/l) and sucrose (3 - 10 %). Regenerating cultures were maintained under a 16-h light photoperiod at 24 ± 1_C.

Source of Information: Plant Tissue Cult. 12(2) : 139-145, 2002 (December)

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

In the present experiment it is observed that the regeneration potential of callus decreased with the age of callus.The observed regeneration in medium with 1 mg/l dicomba. Sucrose plays an important role for cormel formation in vitro in Gladiolus. We observed highest number of cormel formation in medium with 6 % sucrose. The protocol described here is promising for propagation of disease free Gladiolus primulinus cultiver and screening of somaclonal variants. Moreover, it also provides with a source of cells for genetic transformation. The shoots produced roots in half strength of MS with 2.0 mg/l IBA and 6 % sucrose. Rooted shoots produced cormels at the base of shoots in the same nutrient medium. About 80 - 90 % cormels germinated when sown in the field and 20 % of them blossomed in the first year.

Knowledge Management: Journal