Improving propagation of the rare plant Huperzia squarrosa using cuttings and in vitro techniques

Abstract

The tassel fern, Huperzia squarrosa, is a rare and medicinally valuable plant known for containing Huperzine A. It propagates naturally through spores, rhizomes, cuttings, and clump division, but with a slow multiplication rate. This study aimed to optimize propagation conditions for H. squarrosa using stem cuttings and in vitro culture techniques to support its preservation and development. Apical and stem cuttings were treated with varying concentrations of indole-3-butyric acid (IBA) and naphthaleneacetic acid (NAA) before being planted in a substrate of coir dust, charcoal dust, and burnt rice husk (3:2:2). Apical cuttings treated with 1500 ppm IBA for 30 min showed the highest rooting success, identifying this method as optimal for propagation. Additionally, surface sterilization with a 40% bleach solution, followed by antibiotic treatment, achieved a 73.8% clean sample rate. In vitro culturing on ¼ MS (Murashige and Skoog) medium resulted in 70% survival and 55% rooting after 60 days. The highest callus formation rate (13.3%) was achieved with 0.01 mg/L IBA and 0.3 mg/L Kinetin, while the addition of 3 mg/L Glutamine did not significantly enhance callus induction. Ongoing research focuses on enhancing complete plant regeneration and improving the efficiency of in vitro propagation for H. squarrosa.