Development of an Efficient Plant Regeneration System of Field Mustard (Brassica campestris)

Abstract

Aims: The present study was conducted with a view to develop an efficient protocol for high frequency plant regeneration of Brassica campestris for further crop improvement program by biotechnological manipulation and to optimize this system for regeneration of a number of             B. campestris genotypes. Study Design: Completely Randomized Design. Place and Duration of Study: This experiment was carried out in the Genetic Engineering Laboratory of the Department of Genetics and Plant Breeding, Sylhet Agricultural University, Bangladesh during the period of July 2013 to June 2014. Methodology: Cotyledon and hypocotyl explants of B. campestris cv. BARI sarisha-12 were cultured on MS medium supplemented with different concentrations of 6-Benzylaminopurine (BAP) and α-Naphthaleneacetic acid (NAA) for callus initiation and shoot regeneration. Later on subsequent subculturing is done for shoot elongation and multiplication. MS medium supplemented with various concentrations of NAA were used for root formation. Results: From a total of 15 different combinations of BAP and NAA tested, the combination of 1.0 mg L-1 BAP and 0.5 mg L-1 NAA gave the highest frequency of callus initiation (94.44%) as well as shoot regeneration (63.89%) in case of cotyledon explants where as hypocotyl explants showed 47.62% callus initiation and 19.04% shoot regeneration frequency. Four days old cotyledon explants showed the highest shoot regeneration frequency (72.22%) and higher number of shoots per explant (3.94) than those from older seedling. The shoot regeneration frequency markedly enhanced to 83.33% by the addition of 2.0 mg L-1 AgNO3 to the MS medium supplemented with 1.0 mg L-1 BAP, 0.5 mg L-1 NAA and this combination also showed the maximum number of shoots per explant (6.86). Shoot regeneration potentiality of five B. campestris genotypes were investigated and indicated that this system would be widely applicable to all the genotypes. The regenerated shoots were easily rooted on MS medium supplemented with 0.2 mg L-1 NAA and the whole plants were transferred to pot soils and grown to maturity. Conclusion: MS medium supplemented with 1.0 mg L-1 BAP, 0.5 mg L-1 NAA and 2.0 mg L-1 AgNO3 is more efficient for multiple shoot regeneration by using cotyledon explants and it may be utilized for In vitro improvement program of B. campestris.