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Conservation of Mahonia leschenaultii was achieved through multiple axillary shoot formation in single node cultures, in vitro and ex vitro rooting of the shoots, and successful reintroduction and establishment of the rooted plants in the forests of Palani hills in Southern India. A preliminary experiment on the culture of young nodal explants (1.0-1.5 cm) from field-grown plants in different basal nutrient media revealed high frequency bud beak in Schenk and Hildebrandt (SH) medium followed by Woody Plant medium and Murashige Skoog medium in 4 weeks. Subsequent culture of both shoot tips (0.5-0.8 cm) and nodes for 6 weeks in SH medium supplemented with various concentration of cytokinins resulted in optimal formation of 3.6 shoots in 80% of the nodes against 1.35 shoots in 75% of the shoot tips in the presence of 2.0 and 1.0 mg/l benzyl adenine (BA), respectively. Invariably, the resident apical and axillary meristems grew into a single shoot (1.0-1.8 cm) in 1-2 weeks followed by the characteristic differentiation of additional buds from its base. A synergistic combination of 1.0 mg/l BA and 0.02 mg/l indole-3-acetic acid (IAA) induced the maximum number (5.9) of axillary shoot formation which were relatively high (75%) when the explant was collected during May-June and the fifth nodes from the top of the growing shoots were used. Repeated subculture of the nodes from shoot cultures at 5-6 week intervals in medium supplemented with reduced concentrations of the growth regulators (0.5 mg/l BA, 0.01 mg/l IAA) through at least 10 passages enabled consistent production of 6-7 shoots per node at 92% success rate without loss of vigour, growth and morphological abnormalities. Shoots of 3-6 cm were rooted in vitro in the presence of 1.0 mg/l indole-3-butyric acid (IBA) and hardening in the mist house at 76-78% and these rooted plants established in a potting medium of river sand and top soil (1:1) under constant mist irrigation. The plants reared in the nursery for 5-8 weeks were successfully reintroduced into the natural forest segment of the Institute's campus (MSL 200 m) at Palode and Vattakanal shola forests of Palani hills (MSL 2500 m) with establishment frequencies of 78.8 and 90.6, respectively, after 18 months. Growth characteristics of the plants (height, number of leaves and branches) reintroduced into native habitat at Vattakanal were better than those at Palode. Genetic fidelity of the 12-month old plants established in the nursery/field was confirmed by analysis of amplified products of genomic DNA obtained with random primers. |
