In vitro conservation and cryopreservation of mature pistachio (Pistacia vera L.) germplasm

Abstract

As genetic erosion of pistachio (Pistacia vera L.) has been occurring in the Mediterranean, Central and West Asia and North Africa, experiments were conducted to conserve two cultivars (‘Atlı’ and ‘Siirt’) of mature pistachio germplasm by assessing both medium- and long-term conservation techniques. In medium-term conservation, our results showed that it was feasible to conserve both cultivars in the form of either microshoots or encapsulated shoot apices up to 12 months at 4°C in the dark. As regards long-term conservation, encapsulation-dehydration and droplet-vitrification techniques were assessed for cryopreservation of cold-hardened and osmoprotected shoot apices of mature ‘Atlı’ cultivar. Among the methods tested, 13.6% of regrowth was achieved with incubation of explants in the droplets of vitrification solution for 150 min at 0°C followed by direct immersion in liquid nitrogen (LN), rapidly thawed and then cultured on Murashige and Skoog’s (MS) medium containing 1 mg L⁻¹ BA and 0.5 mg L⁻¹ GA_3. The developed droplet-vitrification technique appeared as a promising procedure for long-term preservation of shoot apices of mature pistachio germplasm. Moreover, assesment of genetic fidelity by Random Amplified Polymorphic DNA analysis (RAPD) revealed out high levels of genetic stability between donor plant and cryopreserved plants (similarity indexes between 0.959 and 0.973) after they were subcultured for at least 3 months. The detected low level of genetic instability could be due to the toxic effect of PVS2 and regeneration phase. The optimized conservation techniques, especially slow growth storage, could be applied to preserve other Pistacia species.