PLANT-REGENERATION FROM EXPLANT AND PROTOPLAST-DERIVED CALLUSES OF MEDICAGO-LITTORALIS

Plant regeneration from explant and protoplast derived callus has been achieved in Medicago littoralis cv. Harbinger 1886, an annual legume resistant to the fungus Pseudopeziza medicaginis. Callus was induced f rom different tissue explants and the fastest growth rate was observed for hypocotyls in B5 medium with 2 mg l(-1) 2,4-dichlorophenoxyacetic acid and 0.5 mg l(-1) N-6-benzyladenine. Protoplasts were isolated fr om cotyledons and leaves of sterile plants and from callus; the first two kinds of protoplasts showed a plating efficiency of 5.6% and 5%, r espectively, when embedded in agarose. Plant regeneration occurred on media containing N-6-Delta(2)isopentenyl-adenine combined with indole- 3-acetic acid or 1,2-benzisoxazole-3-acetic acid, and on media with N- 6-benzyladenine plus alpha-naphtaleneacetic acid; a cytokinin/auxin ra tio higher than 1 induced embryos while a ratio around 1 stimulated sh oot formation. Embryo development and rooting of shoots were performed in RL medium without growth regulators.