Micropropagation, callus and root culture of Phyllanthus urinaria (Euphorbiaceae)

Efficient micropropagation, callus culture and root culture protocols were developed for the medicinal plant Phyllanthus urinaria (Euphorbiaceae) using single node explants. Maximum multiplication (16-20 shoots per explant) was achieved on Murashige and Skoog media supplemented with 5.0 muM kinetin. Murashige and Skoog and Anderson Rhododendron media promoted significant shoot culture growth in terms of numbers of shoots and nodes produced per explant. Rooting was achieved with 93-100% of the microshoots on Murashige and Skoog medium without growth regulators, although 1.25-5.0 muM alpha-naphthaleneacetic acid significantly increased the number of roots per explant. Regenerated plants were successfully acclimatized and 91% of plantlets survived under ex vitro conditions. Flowering was observed on micropropagated plants after 3-4 weeks of acclimatization. High frequency callus initiation and growth was achieved when single node explants were inoculated in the horizontal position on Murashige and Skoog medium supplemented with 5.0 muM indole-3-butyric acid. Other auxins such as 2,4-dichlorophenoxyacetic acid and alpha-naphthaleneacetic acid promoted moderate callus fresh weight increase, when used separately. Root cultures were successfully established on Murashige and Skoog medium containing 1.1 muM alpha-naphthaleneacetic acid. The optimized micropropagation, callus culture and root culture protocols offer the possibility to use cell/root culture techniques for vegetative propagation and secondary metabolism studies.