bioRxiv2025
In the course of this study, a protocol for in vitro deposition of a rare plant for the Rostov region, Thymus calcareus, was developed. Before sterilization, the seeds must be subjected to weekly extreme stratification (-18{degrees}C). An effective method for sterilizing primary explants is a mixture of 96% ethanol and 3% hydrogen peroxide in a 1:1 ratio with an exposure time of 7 minutes, followed by three rinses with distilled water. The effective environment for seed initiation is 1/2 MS. The optimal nutrient medium for clonal reproduction of T.calcareus has been determined: 1/2 QL. During long-term cultivation of T.calcareus on a nutrient medium of 1/2 QL, 100% rhizogenesis of plants was observed. The effect of nutrient media, growth regulators and their concentrations on the multiplication and rhizogenesis of T.calcareus explants in vitro has been studied: for multiplication, it is most effective to use a 1/2 QL nutrient medium with the addition of low concentrations of KIN; for rhizogenesis, it is recommended to use QL nutrient media with low concentrations of IAA. The degree of change in the influence of environmental factors, growth regulators and their concentration on T.calcareus in vitro and their change over time on the multiplication rates were determined: the nutrient medium (Ffact = 4.72), the type of growth regulator (Ffact = 7.82) and its concentration (Ffact = 4.71) have a significant effect on the multiplication coefficient of explants. The effect of the factors remains almost unchanged for the mineral base and growth regulator, while the concentration of the growth stimulant increases significantly.; A significant effect on the shoot elongation index was found in the growth regulator (Ffact = 10.39) and its concentration (Ffact = 5.51), the influence of both factors increases