Efficient in vitro propagation and conservation of the endangered medicinal plant Hedyotis sithiravaraiensis
摘要
Hedyotis sithiravaraiensis is an endangered medicinal plant species in Southern India. Due to limited propagation, overharvesting, and habitat loss, the species faces conservation challenges. This study established an efficient in vitro propagation system for the sustainable conservation and utilization of H. sithiravaraiensis. Additionally, it reports on callus induction, shoot organogenesis, and plant regeneration using young leaves of this species. Gamborg B5 media, augmented with various plant growth regulators, was evaluated for their effectiveness in promoting in vitro propagation of H. sithiravaraiensis (Senthumbai). Optimal callus induction was observed on Gamborg B5 media with the addition of 1.0 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D), resulting in a 97.30% induction rate and 3.11 g fresh weight (FW) of callus obtained after 6 weeks. The maximum shoot proliferation frequency (100%) and the maximum shoot number (17.25) were obtained on the B5 media with 1.0 mg/L Kinetin (KN) and 0.2 mg/L naphthalene acetic acid (NAA). Additionally, Half-strength B5 media containing 1.0 mg/L indole-3-butyric acid (IBA) was identified as the best rooting media, resulting in an average of 11.0 roots per shoot and a 100% rooting success after 4 weeks. In vitro cultures were acclimatized by gradual exposure to natural conditions with 80% humidity and reduced photorespiration, resulting in robust leaf development and a 100% survival rate after 6 weeks. Genetic fidelity was further evaluated using two markers, RAPD and ISSR, which produced monomorphic banding patterns in the regenerants, indicating the absence of somaclonal variation and confirming their genetic similarity to the mother plant. The efficient indirect organogenesis method established in this study may significantly contribute to the high clonal propagation and conservation of H. sithiravaraiensis, a medicinally important plant.