Regeneration of Medicago truncatula from tissue culture: increased somatic embryogenesis using explants from regenerated plants

Abstract

Plant regeneration has been achieved by somatic embryogenesis in Medicago truncatula Gaertn. (barrel medic) c.v. Jemalong, an annual legume species. Regenerated plants were obtained from cultured leaf tissue explants on a four-step modified B5 basal medium. Induction of embryo formation occurred on a medium containing 10 μM NAA and 10 μM BAP, and embryo maturation was promoted after transfer to a medium containing 1 μM NAA and 10 μM BAP. Shoot development, secondary somatic embryogenesis and occasional plantlet development occurred on a subsequent transfer to 0.1 μM NAA and 1 μM BAP. Plantlet formation could also be completed by transfer of well developed shoots to 0.05 μM NAA. A high frequency of primary somatic embryos could only be obtained by using the same culture protocol with tissue from regenerated plants. Explants from regenerated plants showed a large increase in the number of primary embryos per callus and the number of calli producing embryos. Explants from plants derived from the seed of one regenerated plant also showed increased embryo formation. Although high embryo formation rates can be reproducibly obtained from this seed, embryo conversion rates to plants are currently low.