Somatic embryogenesis in Crambe abyssinica Hochst. ex R.E. Fries using seedling explants

Abstract

Somatic embryogenesis in Crambe abyssinica, an important industrial oilseed species, was investigated. Cotyledon, hypocotyl and root explants from 8-day-old seedlings of C. abyssinica cv. Prophet were cultured with levels of 1-naphthaleneacetic acid (NAA) and 2,4-dichlorophenoxyacetic acid (2,4-D) ranging from 2.2 to 39.0 μM, combined with 6-benzyladenine (BA) to achieve an auxin:cytokinin ratio of 20:1, and callus formation assessed. Callus formation frequency for cotyledon and hypocotyl explants was 100% for levels of 2,4-D from 4.5 to 33.9 μM. The response was similar with NAA levels of 13.0 to 39.0 μM. Root explants were less responsive. When calluses were transferred to a medium containing 0.56 μM each of thidiazuron and BA with 1.0 μM indole-3-butyric acid (IBA), somatic embryos were induced. Moreover, embryos were induced from calluses grown on media containing either 11.3 μM 2,4-D or 13.0 μM NAA, or higher. On a medium without plant growth regulators, embryos were induced but at a much lower frequency. For all three explants, 22.6 μM 2,4-D and 26.0 μM NAA was optimal for embryogenic callus induction. Hypocotyl-derived calluses were superior to cotyledon- and root-derived calluses for embryo induction. The best embryo formation response was with medium containing 5.0–6.0% sucrose. The highest average number of embryos per callus (36) was obtained from hypocotyl calluses from medium with 22.6 μM 2,4-D. Somatic embryos germinated best on half-strength B5 or MS medium with 3% sucrose, and plantlets were successfully established under greenhouse conditions. The results indicate that high levels of auxins are required for the induction of embryogenic calluses from explants of C. abyssinicia, while cytokinins are critical for somatic embryo formation.