High frequency plant regeneration in Indian Siris via cyclic somatic embryogenesis with biochemical, histological and SEM investigations

Abstract

The present work describes an efficient system for cyclic somatic embryogenesis along with biochemical determination of antioxidative enzymes at characteristic stages of embryonic development in Albizia lebbeck. Highest embryogenic callii induction with maximum development of primary somatic embryoids (70.3±0.88%) was achieved on Woody Plant Medium (WPM, 1981) supplemented with 12.5μM kinetin (KN), whereas enhanced maturation (53.67±0.88%) of primary somatic embryoids and the induction of cyclic somatic embryoids occurred on Murashige and Skoog’s medium (MS, 1962) containing 5.0μM meta-Topolin (mT), 2.5μM α-naphthaleneacetic acid (NAA) and 75.0μM Glutamine (Gt). 30g/L sucrose and low pH (5.8) were found to be most conducive to both primary and secondary somatic embryogenesis. Maximum germination and conversion of primary and secondary somatic embryoids into plantlets were achieved on half-strength MS medium supplemented with 1.0μM gibberellic acid (GA3). About 50% of somatic embryoid derived plantlets were successfully acclimatised to ex vitro conditions and showed normal phenotypes. Regarding oxidative events, upregulated levels of superoxide dismutase (SOD), ascorbate peroxidase (APX) and glutathione reductase (GR) were recorded during early stages of embryoid development. In contrast, decreased catalase (CAT) activities were detected at initial stages of embryoid development suggesting higher intracellular H2O2 levels in embryogenic cells. Moreover, histological analysis in conjuction with scanning electron microscopic studies was carried out to confirm the ontogeny, morphology and development of somatic embryoids.