Abstract
International trade in medicinal plants is a major force in the current global economy and the demand of natural health products is increasing in developing and industrialized nations. Unfortunately, one third of all plant species and medicinal plants in particular, are under threat of extinction due to indiscriminate harvest, diseases, and reduced habitats. In vitro culture and cryopreservation techniques can ensure safety of these species and have potential applications in sustainable production and use of plant based medicines. The objective of this study was to develop an efficient cryopreservation protocol for in vitro maintained lines of medicinal plants using St John's Wort and tobacco as experimental systems. Shoot tips of in vitro-grown plantlets were successfully cryopreserved in liquid nitrogen at -196°C under controlled environment conditions following melatonin treatment and cold acclimation (CA) with either vitrification or encapsulation-vitrification methods. The explants showed optimal re-growth following cryopreservation when treated with the Plant Vitrification Solution #2 (PVS2) for 20 min on ice. Addition of the antioxidant melatonin (0.1-0.5 µM) added to both pre-culture and re-growth media significantly improved the recovery of plants with 80 to 100% re-growth of cryopreserved shoots. The melatonin based protocol for improved recovery of cryopreserved tissue may serve as an important tool for long-term storage and security of germplasm of medicinal plant species.
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