Assessment of polysomaty, embryo formation and regeneration in liquid media for various species of diploid annual Medicago

To avoid polyploidy in regenerants the source of explant material should be monosomatic. Therefore, the leaf and petiole tissue of five diploid Medica go species (Medicago ciliaris, Medicago murex, Medicago orbicularis, Medica go polymorpha and Medicago tnuncatula cv. Jemalong, and the ecotype R108-1) was assessed for polysomaty by flow cytometry. For the species studied the frequency of 2C nuclei was about 90% in leaves compared with that in petio les. Embryos were readily formed from tissue of leaves in liquid media cont aining 1 mg 1(-1) or 4 mg 1(-1) dichlorophenoxyacetic acid (2,4-D). For emb ryo development two procedures were tested - prolonged use of induction med ium and treatment with polyethylene glycol Mw 6000 (PEG). The highly regene rable genotypes M, truncatula cv. Jemalong and R108-1 showed efficient conv ersion of embryos after maturation in liquid medium. The regenerated plants were diploid and with normal phenotype. (C) 2001 Elsevier Science Ireland Ltd. All rights reserved.