Shoot tip cryopreservation by vitrification in Kaempferia galanga L. An endangered, overexploited medicinal plant in Tropical Asia

Abstract

Shoot tips from the established in vitro shoot cultures of Kaempferia galanga L., an endangered medicinal plant were successfully cryopreserved using the vitrification procedure. Effect of concentration of sucrose for preculture, vitrification treatment with PVS2(Plant vitrification solution 2) and recovery medium on cryopreservation of shoot tip meristems has been analyzed. Overnight preculturing of the dissected shoot tips in MS medium containing 0.4 M sucrose, osmoprotection with loading solution for 20 minutes, dehydration with PVS2 for 20 minutes at 0 oC were found to be optimum among the various treatments tested. The cryoprotected shoot tips upon rapid freezing in LN (liquid nitrogen) followed by rapid thawing produced 50-60% survival and 30-40% regeneration rates. Incorporation of GA3 (giberellic acid) in the post-thaw culture medium was essential to induce shoot emergence in the initial phase. Subculturing of the recovered shoot tips in MS medium supplemented with 1.0 mgl -1 BA (6-benzyl adenine) and 0.5 mgl -1 NAA (α-naphthaleneacetic acid) resulted plantlet production. The regenerated shoots grew to mature plants were free of any morphogenetic variation upon field transfer.