In vitro culture and micropropagation of the Baetic-Moroccan endemic plant Lapiedra martinezii Lag. (Amaryllidaceae)

Abstract

Lapiedra martinezii Lag. is a potential medicinal and ornamental plant facing conservation challenges. Thus, this study was focused on determining the conditions for culture initiation and propagation using in vitro techniques. The optimal sterilization procedure combined thermotherapy at 54°C for 60 min and immersion in 7% (w/v) Ca(ClO)2 solution for 20 min. The most suitable medium to initiate bulb scales cultures was Gamborg B5 medium containing 500 mg L−1 casein, 2 mg L−1 adenine, 10 mg L−1 glutathione and 10 g L−1 sucrose. The most productive multiplication medium tested was Murashige and Skoog medium containing 30 g L−1 sucrose, 4.0 mg L−1 6-benzylaminopurine, and 0.12 mg L−1 1-naphtaleneacetic acid. Most plants developed in vitro rooted spontaneously in the multiplication phase. The vast majority of the plants (89%) were successfully transferred to ex vitro conditions, and 100% survived over 1 yr of cultivation outdoors. Sucrose at a concentration of 60 g L−1 was the most effective treatment to increase the biomass of bulblets. High auxin/cytokinin ratios produced the highest callus induction efficiency. The vast majority of callus developed in dark conditions, but none regenerated in the combinations of growth regulators previously tested. The plants obtained by micropropagation did not show significant differences in morphometric traits compared with the wild specimens, which supported the stability of the materials produced in vitro. This is the first report on cell cultures and micropropagation of L. martinezii, and the results can be applied to other Amaryllidaceae for industrial or conservation purposes.