An Effective Protocol for the Production of Primary and Secondary Somatic Embryos in Cauliflower (Brassica Oleraceae Var. Botrytis)

An in vitro regeneration protocol based on the use of root-derived embryogenic callus tissue was established. Three different sizes of explants were tested in order to determine the optimal size of explants for somatic embryo production. The best formation of somatic embryos was on explant size 600-1000^m achieved using 90sec duration of blending. Kinetin played an important role in somatic embryo induction, development and maturation. Embryo maturation occurred on the medium with the lowest concentration of kinetin (0.5mg L-1). Normal secondary somatic embryos (SSEs) were induced directly on hypocotyls of primary somatic when cultured on semi solid MS basal medium supplemented with 30g L-1 sucrose in the absence of growth regulators. SSEs completed full development in the same medium. Cyclic secondary somatic embryogenesis was a good source of embryogenic material, which can be used for increasing the efficiency of plantlet regeneration in cauliflower. After 60 days of culture, it was demonstrated that using activated charcoal in the culture medium had no significant effect on SSEs yield and moreover, it had negative impacts on the morphological shape of embryos as most of SSEs were abnormal with split collar cotyledons and were smaller in size. The effective protocol for somatic embryo production designed in this study could have important applications in the field of cauliflower micropropagation and breeding systems.