A suitable protocol for in vitro tuber production using non-dormant tubers of Gloriosa superba L. on Murashige and Skoog (MS) medium without addition of plant growth regulators is reported in the present study. Among the different basal media tested MS medium was found to be suitable for induction and development of secondary tubers; one in vitro tuber per explant was obtained after 6 weeks and 3 tubers per explant after 12 weeks of culture. These tubers produced healthy green shoots that rooted on basal medium. Best rooting was noted in half strength (half organics and inorganics) MS medium with one-fifth nitrates. © 2007 Elsevier B.V. All rights reserved.