The conservation of pteridophytes presents a demanding challenge because many species in the world are thought to be threatened with extinction, as they are very sensitive to environmental disturbance. Ex situ actions provide an important conservation strategy, so the Germplasm Bank of Tuscia Botanic Garden, with the Herbarium UTV (Tuscia University, Viterbo–Italy), has undertaken a project for the conservation of threatened pteridophytes of the Italian flora, like Osmunda regalis, the Royal Fern, a species with chlorophyllous spores that is declining in Italy because it is linked to vulnerable habitats. As a part of the project, this work presents first results of in vitro reproduction of the Royal Fern using spores from exsiccata (UTV samples collected and dried in 1989 and 2001). Our results also highlight the value of herbarium specimens in biodiversity conservation, providing a useful method to reproduce species that are threatened or extinct in the wild, at least at a local level, so as to plan eventual reintroductions using the native germplasm.