Enhanced bud and bulblet regeneration from bulbs of Nerine sarniensis cultured in vitro

Abstract

Nerine (Nerine sarniensis) cv. Salmon Supreme in vitro-grown bulblets, 7-9 mm in diameter, were cut in half longitudinally and used for adventitious bud initiation following dissection of the roots and two-thirds of the upper part of the bulblets. The terminal apex was injured with a hot, sterile microscope dissecting needle. The highest number of buds formed (seven to nine buds per halved bulblet) on a semi-solid Murashige and Skoog (MS) basal salts medium supplemented with 3% sucrose and either 1 microM 6-benzylaminopurine (BA) and 1 microM alpha-naphthaleneacetic acid (NAA) or 0.5 microM BA and 0.1 microM NAA. Bulblet halves were cultured in the dark for 11-13 weeks with one subculture after 6 weeks. Anatomical studies indicated that the initiation of adventitious buds on the abaxial side of the inner scales of the halved bulblet was adjacent to the basal plate and started from the leaf primordium and a meristematic bulge. Buds developed directly into small bulblets after they were transferred to semi-solid MS basal salts medium supplemented with 6% sucrose, 10 microM indole-3-butyric acid (IBA) and 0.25% activated charcoal. Small bulblets cultured in liquid MS medium supplemented with additional KH(2)PO(4 )(170 mg l(-1)), 6% sucrose and 0.1 microM NAA under a 16/8-h (light/dark) photoperiod for 8 weeks grew into larger bulbs faster than those cultured on semi-solid medium. The bulbs were rooted on a semi-solid medium after 4 weeks and then transferred to the soil. As many as 18 bulblets developed and rooted from one in vitro-grown bulb after 25-27 weeks.