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Callus Derived Regeneration of Some Selected Brassica Genotypes

S. K. Biswas
Jute Agricultural Experimental Station, Bangladesh Jute Research Institute (BJRI), Manikganj, Bangladesh

M. Z. Tareq
Jute Agricultural Experimental Station, Bangladesh Jute Research Institute (BJRI), Manikganj, Bangladesh

S. Ahmmed
Pest Management Division, BJRI, Dhaka, Bangladesh

A. B. M. Z. Hoque
Genetic Resource & Seed Division, BJRI, Dhaka, Bangladesh

M. T. Rahman
Jute Seed Production and Research Division, BJRI, Dinajpur, Bangladesh

Abstract/ Executive Summary:

The experiment was conducted to observe the callus induction ability of Brassica species. Plantlets were regenerated from cotyledon and stem explants of Brassica napus, Brassica campestris and Brassica juncea through direct organogenesis. The experiments were conducted in a Completely Randomized Design (CRD) with 4 replications. The highest frequency of callus formation was recorded in MS containing 2.0 mgl^-1 BAP, 0.5 mgl^-1 NAA and 2.0 mgl^-1 AgNO₃ in both stem and cotyledon explants. Among these explants, stem was found to be better responsive in callus induction than cotyledon. Among the genotypes used, BINA Sarisha-4 induced the highest percentage (100.00%) of callus from stem explants which was followed by BINA Sarisha-5 (100.00%) and Sampad (83.35%). On the other hand, BINA Sarisha-4 induced callus from 91.67% cotyledon explants, followed by BINA Sarisha-5 (75.00%) and Sampad (66.67%). Similarly, the highest percentage of shoot regeneration (58.34%) from stem explants of BINA Sarisha-4 was observed in MS medium supplemented with combination of hormone and silver nitrate concentrations. The highest percentage of root induction was 66.67 and 58.33% in plantlets derived from stem and cotyledon explants, respectively in ½ MS medium supplemented with 2.0 mgl^-1 IBA and 0.5 mgl^-1 of NAA. The highest survival rate was found after acclimatization of plants derived from stem (77.78%) and cotyledon (64.28%) explants of BINA Sarisha-4 in pot and 64.33 and 55.55%, respectively in field.The experiment was conducted to observe the callus induction ability of Brassica species. Plantlets were regenerated from cotyledon and stem explants of Brassica napus, Brassica campestris and Brassica juncea through direct organogenesis. The experiments were conducted in a Completely Randomized Design (CRD) with 4 replications. The highest frequency of callus formation was recorded in MS containing 2.0 mgl^-1 BAP, 0.5 mgl^-1 NAA and 2.0 mgl^-1 AgNO₃ in both stem and cotyledon explants. Among these explants, stem was found to be better responsive in callus induction than cotyledon. Among the genotypes used, BINA Sarisha-4 induced the highest percentage (100.00%) of callus from stem explants which was followed by BINA Sarisha-5 (100.00%) and Sampad (83.35%). On the other hand, BINA Sarisha-4 induced callus from 91.67% cotyledon explants, followed by BINA Sarisha-5 (75.00%) and Sampad (66.67%). Similarly, the highest percentage of shoot regeneration (58.34%) from stem explants of BINA Sarisha-4 was observed in MS medium supplemented with combination of hormone and silver nitrate concentrations. The highest percentage of root induction was 66.67 and 58.33% in plantlets derived from stem and cotyledon explants, respectively in ½ MS medium supplemented with 2.0 mgl^-1 IBA and 0.5 mgl^-1 of NAA. The highest survival rate was found after acclimatization of plants derived from stem (77.78%) and cotyledon (64.28%) explants of BINA Sarisha-4 in pot and 64.33 and 55.55%, respectively in field.

Keywords: In vitro, Organogenesis, Cotyledon, Stem explants, Mustard

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

Start Date: 00-00-2007

End Date: 00-00-2008

Program Area: Variety and Species

Commodity: Mustard

Objectives:

To study the in vitro regeneration potentiality of four selected Brassica genotypes.

Methodology:

Plant materials- Four different varieties of Brassica spp. such as BINA Sarisha-4 and BINA Sarisha-5 of Brassica napus, ‘Sampad’ of Brassica campestris and ‘Shambal’ of Brassica juncea were used in this study. The seed materials were collected both from Bangladesh Institute of Nuclear Agriculture (BINA) and Bangladesh Agricultural University (BAU), Mymensingh. Present research activities were conducted at the Tissue Culture Laboratory of the Department of Genetics and Plant Breeding, BAU, during the year 2007-2008. Culture medium and explants used- Mature seeds of Brassica napus, Brassica juncea and Brassica campestris were surface sterilized in a sequential manner with 70% ethyl alcohol for five minutes, 0.1% HgCl₂ with 1-2 drop Tween-20 for five minutes followed by treatment with sodium hypochlorite (2.5% Chlorine) for five minutes and subsequently rinsed four to five times with sterile distilled water. Sterilized seeds were germinated aseptically on half-strength MS basal medium. MS medium contained inorganic salts of MS medium, vitamins, 20 gml^-1 sucrose and 6 gm agarose (Difco-brand Bacto Agar). The pH of all culture medium used in this study was adjusted to 5.8 before autoclaving at 121^oC for 30 minutes. Cotyledons with 1-2 mm petiole and stem segments of 2 to 3 mm in length were cut from four-day old germinated seedlings. Cotyledons were placed upright with cut ends embedded in the culture medium and stem segments were cultured horizontally and pressed down gently into the medium. Four explants were cultured per Petridish. The cultured plates were sealed using micro porous tape and incubated at 22 ± 2°C using 16-hr. day length (2000Lux). Color, nature and abundance of callus were observed visually after three weeks of incubation and graded accordingly. The color graded the marks as 3 for green, 2 for creamy and 1 for yellow color; in case of nature of callus 3 for compact, 2 for friable and 1 for looses in texture and in case of abundance of callus 3 for plenty, 2 for moderate and 1 for poor amount. Callus induction and shoot regeneration from stem segment and cotyledon of four Brassica genotypes were cultured on MS medium supplemented with different concentrations of BAP (1 and 2 mgl^-1) and NAA (0.5 and 1.0 mgl^-1) with constant concentrations of AgNO₃ (2.0 mgl^-1) in order to induce shoot from unorganized calli. After two weeks shoot initiation was visible and well developed shoots (3-4 cm long) were cultured on root induction medium (half- strength MS) supplemented with different concentrations of IBA (1, 2 and 3 mgl^-1) with constant concentration of NAA (0.5 mgl^-1). After 12 weeks culture the number of shoots per explant was recorded. Most of the shoots become rooted on rooting medium. Rooted shoots were planted in a mixer soil containing 25% garden soil+ 50% sand+ 25% cow dung in plastic pots and kept in a lath house for hardening before transferring them to field. The experiments were conducted following Completely Randomized Design (CRD) with 4 replications. Statistical analysis- The data collected on different characters were analysis statistically to find out the analyses of variance and means were compared by the Duncan’s Multiple Range Test (DMRT).

Source of Information: The Agriculturists 15(2): 01-10 (2017)

Article Link (Online Journal):

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

From the above study it can be concluded that stem segment was better than cotyledon in callus formation, shoot regeneration, root initiation and establishment of plantlets in soil. Among the genotypes, BINA Sarisha-4 performed the best in all cases.

Knowledge Management: Journal