In vitro selection and molecular characterization of salt tolerant canola plantlets

ABSTRACT

In this study, a system for in vitro selection of salt tolerant canola clones [plantlets] was developed. The developed system relies on somaclonal variation and the high regenerative capacity of hypocotyl explants of the Egyptian canola cultivar Serw 4 via direct and indirect shoot organogenesis. The optimized regeneration system involves callus formation from hypocotyl explants of 4 day-old seedlings on MS medium supplemented with 1.5 mg/l 2,4-D and shoot proliferation from proliferated microcalli colonies on MS + 4.0 mg/l BA. Shoot organogenesis frequecny ranged from 35 to 40%. This system was used for in vitro selection of salt tolerant plantlets from hypocotyl explants of seedlings germinated on salt containing medium. Under selection pressure, shoot organogenesis frequencey dropped to 5% for selected cell lines and zero for non-selected cell lines onto 0.3% NaCl. The biochemical and molecular characterization of the selected and non-selected clones were performed using proline accumulation, total protein content, SDS-PAGE and RAPD analysis. The salt-tolerant plantlets were characterized by improved growth on saline media, rapid accumulation of proline, high content of total protein, synthesis of new set of high molecular weight proteins [38 and 94 KD] and unique RAPD banding profiles. The criteria of the selected clones outlined above may be a part of biochemical make up performed by these canola clones to tolerate extreme salt stress