High-frequency Organogenesis from Direct Seed Culture in Lathyrus

Abstract

Culture conditions were developed for inducing a high frequency of direct shoot morphogenesis and whole plant regeneration in cultures of intact seedlings of Lathyrus cicera L., L. ochrus (L.) DC., L. sativus L., and L. tingitanus L. The procedure of shoot regeneration involved culturing of whole, mature seeds on MS medium containing cytokinins, or thidiazuron (TDZ), a substituted phenylurea with cytokinin-like activity. Differentiation of shoots occurred without an intervening callus phase from the cotyledonary node and surrounding tissues of the intact seedlings developed from seeds germinated on media containing kinetin, BAP or TDZ. An average of 19·0, 15·8, 28·8 and 43·0 shoots were regenerated at optimal concentrations of 50·0, 50·0 and 80·0 μM BAP in L. cicera, L. tingitanus, L. ochrus and L. sativus, respectively. TDZ enhanced the shoot formation at the concentration of 10 μM to an average of 33·1 and 33·7 shoots per seedling in L. cicera and L. tingitanus and at 50 μM, to 57·4 shoots per seedling in L. sativus. Regenerated shoots developed roots on a modified MS medium containing 2·5 μM NAA and the surviving plantlets were transferred to soil. Histological studies revealed de novo formation of shoot buds from the epidermal or subepidermal cells of the basal and nodal region of multiple epicotyls. Several meristematic centres consisting of actively-dividing cells developed in the subepidermal cell layer of the nodal tissue and adjacent areas within 7 d of seed culture on a cytokinin- or TDZ-supplemented medium. The pattern of the development of these meristematic centres and shoot development was similar in all four species.