Biotechnological advances for preserving the endangered Sicilian fir Abies nebrodensis

Somatic embryogenesis (SE) stands as a pioneering biotechnological technique employed for plant cloning, with vast applications in propagation, conservation, and cloning of diverse plant species. Abies nebrodensis, an endangered species exclusive to the Madonie Park in Sicily, Italy, has been a focal point of conservation efforts due to its critically low population of 30 adult individuals worldwide. The LIFE European program initiated the Life4Fir project to safeguard this species by employing innovative biotechnological tools. In response to these challenges, the present work outlines the exclusive protocol for SE in the Sicilian fir Abies nebrodensis. Cones from identified Abies adult trees were collected, and experiments were conducted to initiate callus from both zygotic immature and mature embryos. Successful results were achieved with SH medium supplemented with 1 mg L-1 of 6-benzylaminopurine (BAP). Furthermore, development of somatic embryos occurred on SH basal salt medium with abscisic acid (ABA) and polyethylene glycol (PEG-4000). The study highlights the impact of culture medium, PGR type, donor tree origin, and seed storage duration on SE outcomes. Moreover, cryopreservation, preserving tissues at -196°C, has been successfully applied, providing effective protocols for preserving pollen, embryos, and embryogenic callus lines. This establishes the groundwork for a cryo-bank for A. nebrodensis, enhancing the prospects for large-scale propagation and long-term preservation of this endangered species.