A procedure for indirect shoot organogenesis of Polianthes tuberosa L. and analysis of genetic stability using ISSR markers in regenerated plants

Efficient in vitro regeneration system was well-established in Polianthes tuberosa L., an attractive ornamental and aromatic plant of horticultural importance. Various explants (Crown explants, leaves, petals, flowering stems and bulb scales) and different plant growth regulators (PGRs) were evaluated for both callus induction and shoot regeneration. Results showed that the maximum callus formation was obtained in crown explants using Murashige and Skoog (MS) medium supplemented by 1.95 mu M 2,4,5-Trichlorophenoxyacetic acid (2,4,5-T). The highest rate of shoot regeneration frequency was obtained on MS medium enriched with 2.26 mu M thidiazuron (TDZ) + 250 mgl(-1) proline from crown callus. The regenerated shoots were elongated on MS medium supplemented with 2.22 mu M 6-benzylaminopurine (BAP). All shoots were successfully rooted in MS medium containing 9.89 mu M indole-3-butyric acid (IBA) within a 15-day under a 16/8 h photoperiod. Plantlets were successfully hardened and adapted in the greenhouse with a survival rate of 100% and demonstrated normal growth. Flow cytometry and Inter Simple Sequence Repeats (ISSR) markers analysis indicated that there was no variation between mother and regenerated plants. The established regeneration protocol would be very useful for genetic engineering of P. tuberosa L. leading to the production of new flower colour and shape, pest and disease resistance and fragrance modification.