Using Stress Factors for Storage of Withania somnifera L. Hairy Roots without Passages

Hairy root cultures are widely used both for fundamental and practical purposes as a source of valuable secondary metabolites. Hairy roots require frequent passages to fresh nutrient medium due to their rapid growth. Cryopreservation of the most productive hairy root lines for a long time (some years) is a resource-consuming task. In this regard, it is especially important to develop alternative methods for short-term (few months) storage of hairy roots without the loss of their properties and viability. In this study hairy roots of Withania somnifera L. were used as an experimental object to test different stressful conditions for short-term preservation. Effects of increased content of mannitol and NaCl in the medium, oxygen and sucrose deficiency and cold stress on the root growth parameters were evaluated. Nutrient medium with high content of NaCl and reduced concentration of sucrose was the most suitable for the short-term preservation of hairy roots. All cultures survived in the presence of either 300 mM NaCl or 1.875 g/L sucrose for three months. Standard activity of catalases and peroxidases, malondialdehyde content and total antioxidant capacities were completely restored in the cultures after they were returned to normal conditions. Moreover, these hairy roots maintained the same growth rate they had before the treatment. According to these results, we propose new, simple and inexpensive methods for long-term storage of hairy roots at room temperature.