Direct somatic embryogenesis, plant regeneration and in vitro flowering in rapid-cycling Brassica napus

Abstract

A simple method to induce somatic embryogenesis from seeds of rapid-cycling Brassica napus is described. Seedlings cultured on Murashige and Skoog (MS) basal medium produced somatic embryos directly on hypocotyls and cotyledons after 2 to 3 subcultures onto the same medium. A low pH of the medium (3.5-5) was more conducive to somatic embryogenesis than a higher pH (6 and 7). Embryogenic potential of the seeds was inversely correlated to seed age: about 41-68% of immature seeds between the ages of 14 and 28 days after pollination (DAP) formed somatic embryos compared to 0-11% of the seeds obtained 29-37 DAP. About 54% of the somatic embryos produced secondary embryos after subculturing onto the same medium. The embryogenic potential of the cultures has been maintained on MS basal medium for 2 years (12 generations) without diminution. Up to 75% of the secondary embryos developed into plantlets on MS medium enriched with 10-6 M zeatin, and 40% of these produced flowers when transferred to an optimised flower-induction medium. Viable seeds were produced in self-pollinated in vitro flowers.