Optimisation of Low-Temperature In Vitro Conservation of Garlic (Allium sativum L.) Germplasm for Biodiversity Preservation

The conservation of garlic (Allium sativum L.) genetic resources is essential to maintain agricultural biodiversity and preserve valuable local cultivars threatened by climate change, pathogen pressure, and replacement by intensive commercial varieties. In vitro slow-growth storage represents an effective strategy for medium-term germplasm conservation because it reduces maintenance requirements while preserving disease-free plant material. This study aimed to optimise low-temperature in vitro conservation conditions for garlic germplasm using different nutrient media, osmotic agents, and plant growth regulators. Garlic plantlets were cultivated on full-strength and half-strength Murashige and Skoog (MS) media supplemented with sorbitol, mannitol, sucrose, chlormequat chloride (CCC), or abscisic acid (ABA). Cultures were maintained under slow-growth conditions at 5 ± 1 °C. Low-temperature cultivation substantially reduced plant growth while maintaining high viability and plant quality. The strongest growth suppression was achieved on MS medium containing 5 mg L⁻¹ ABA, 4% sorbitol, or on half-strength MS medium supplemented with 10% sucrose. After 5 months of storage, plantlets were transferred to recovery media to evaluate their regeneration capacity. The highest regrowth potential was observed in cultures maintained on 10% sucrose, followed by those stored with 4% sorbitol. Although ABA effectively inhibited growth, post-storage recovery was considerably weaker, indicating prolonged physiological stress. The results demonstrate that low-temperature storage at 5°C is the principal factor enabling successful medium-term conservation of garlic germplasm. Among the tested treatments, sucrose and sorbitol provided the most suitable balance between growth suppression and regeneration capacity. The optimised protocol offers a practical approach for preserving garlic genetic resources and supporting ex situ biodiversity conservation programs while reducing labour, space, and maintenance costs associated with germplasm collections.