Micropropagation techniques, germplasm conservation and genetic transformation studies in Dioscorea prazeri

Abstract

Dioscorea prazeri is a valuable medicinal plant belongs to the genera of yams, found newlinechiefly at higher altitudes of North Eastern Himalayas. Diosgenin, the therapeutically newlinesignificant secondary metabolite present in D. prazeri tubers is amongst the 10 most newlineimportant sources of steroids, widely prescribed medicine of plant origin. This research newlineholds immense potential in exploring the bioactive, Diosgenin and sustainable newlinemanagement and utilization of this valuable medicinal yam, threatened and indigenous to newlineIndia. A highly efficient in vitro regeneration of D. prazeri was achieved using nodal newlineexplants and axillary buds on Murashige and Skoog medium containing sucrose, newlinesupplemented with growth regulators. Germplasm conservation technique was newlineestablished using vitrification and encapsulation dehydration method and a combination newlineof these two. This protocol ensures long-term availability of this precious germplasm and newlinecan be successfully exploited for conservation of Dioscorea species and other elite plant newlinespecies. The plantlets had healthy roots and sprouted tubers in vitro, were subsequently newlineacclimatized and successfully established in soil. The regenerated D. prazeri in this study newlinewould be reintroduced to the natural habitat. The method of extraction and newlinechromatographic analysis were standardized to obtain maximum yield of Diosgenin from newlineextracts of D. prazeri. An examination of the genetic fidelity of the in vitro regenerated newlineplants using morphological, molecular and biochemical methods indicated a stable newlinegenome. The gene of interest, 3-hydroxy-3-methyl-glutaryl-CoA reductase 1, was isolated newlinefrom Hevea brasiliensis for genetic transformation studies on D. prazeri. The RNA was newlinereverse transcribed and amplified with gene specific primers to obtain the 1.8 kb newlinetranscript. The sequence of confirmed gene was further genetically engineered into plant newlineexpression vector, introduced into D. prazeri via A. tumefaciens mediated transformation.