In vitro propagation and reintroduction of the endangered renanthera imschootiana rolfe

Abstract

Renanthera imschootiana Rolfe is an endangered tropical epiphytic orchid that is threatened with extinction due to overcollection and the loss of suitable habitats. In vitro propagation is a useful way to mass produce plants for re-establishment in the wild and for commercial propagation. Seeds collected 150 days after pollination (DAP) were the optimum stage for in vitro culture. Seed germination reached 93.1% on quarter-strength MS (i.e., MS containing a quarter of macro- and micronutrients) medium containing 0.5 mg l-1 α-naphthaleneacetic acid (NAA), 20% coconut water (CW), 1.0 g l-1 peptone, 10 g l-1 sucrose and 1.0 g l-1 activated charcoal (AC). Quarter-strength MS medium supplemented with 1.0 mg l-1 BA, 0.5 mg l-1 NAA, 1.0 g l-1 peptone, 10 g l-1 sucrose and 20% CW was suitable for the sub-culture of protocorm-like bodies (PLBs) in which the PLB proliferation ratio was 2.88. Quarter-strength MS medium containing 1.0 mg l-1 NAA, 1.0 g l-1 peptone, 100 g l-1 banana homogenate (BH), and 1.0 g l-1 AC was suitable for plantlet formation and 95.67% of plantlets developed from PLBs within 60 days of culture. Hyponex N016 medium supplemented with 0.5 mg l-1 NAA, 1.0 g l-1 peptone, 20 g l-1 sucrose, 150 g l-1 BH, and 1.0 g l-1 AC was suitable for the in vitro growth of plantlets about 2-cm in height. Plantlets 3-cm in height or taller were transplanted to Chilean sphagnum moss, and 95% of plantlets survived after 60 days in a greenhouse. Three hundred transplanted of seedlings 360-days old were reintroduced into three natural habitats. Highest percentage survival (79.67%) was observed in Yuanjiang Nature Reserve two years after reintroduction, followed by Huolu Mountain forest park (71.33%). This protocol is an efficient means for the large-scale propagation and in vitro and in vivo germplasm conservation of R. imschootiana.