IMPROVEMENT OF PROTOPLAST REGENERATION FROM A RECALCITRANT INBRED LINE OF BRASSICA-OLERACEA - A MORPHOGENETIC ANALYSIS

The regeneration potency of protoplast-derived calli from a regenerati ve and a recalcitrant inbred line of Brassica oleracea was studied. Mi crocallus of the regenerative line consisted of meristematic structure s that generated multiple meristematic nodules on callus proliferation medium K3. After transfer to regeneration medium K3R multiple shoot p rimordia developed. Microcallus of the recalcitrant line also containe d meristematic regions during early culture, but most of them disappea red after 3 weeks on proliferation medium. Accordingly, a low regenera tion rate was found for this genotype. However, when the calli were tr ansferred to regeneration medium after a culture period of 2 weeks on K3, a fourfold increase in regeneration rate was observed. It is concl uded that the recalcitrance of the non-regenerative line is due to the inability to develop shoot primordia after prolonged culture on K3. W e further noticed that the regeneration rate of both lines is strongly affected by the hormonal composition of the callus proliferation medi um, in particular by the presence of 2,4-D. The period of callus proli feration is therefore considered as a critical step in shoot different iation.