Abstract
<jats:p>The article presents the results of a study aimed at optimizing the biotechnology of micro clonal propagation of three varieties of Fragaria × ananassa in order to obtain healthy, virus-free planting ma terial. The novelty of the study lies in the development of an integrated biotechnological approach to obtaining virus-free strawberry planting material using a new synergistic combination of metabolically active addi tives-proline, adenosine triphosphate (ATP) and ascorbic acid. The proposed strategy reduces stress responses in explants during early cultivation stages, enhances energy metabolism and stabilizes meristem cells, in creasing micropropagation rate and regenerative capacity compared to traditional protocols. This combination was first applied to improve the morphogenetic activity of strawberry explants and demonstrated a sustained 30–45 % increase in reproduction rate depending on genotype. Methodologically, the study included stages of sterilization, regeneration, proliferation, and rooting of apical meristems. It was found that a five-minute treat ment with a 0.1 % sodium hypochlorite (NaClO) solution provides an optimal balance between sterilizations efficiency and tissue survival, reducing the contamination level to 25–35 % with meristem viability of 65–75 %. The addition of ascorbic acid (10 mg/L) prevented phenolic darkening of the meristems. The inclusion of proline and ATP contributed to an increase in the length of micro-rosettes and the reproduction coefficient: in Malwina variety – from 1:11 to 1:16, in Black Prince variety – from 1:7 to 1:11, and in the Sabrina variety – from 1:3 to 1:4. Molecular biological analysis using RT-PCR confirmed the absence of SVBV, SCV, SMoV, SPaV and BPYV viruses, indicating a high degree of recovery of the material obtained. In conclusion, it has been established that the proposed microclonal propagation protocol increases the regenerative capacity of meristems, accelerates the production of virus-free plants, and is recommended for industrial production of strawberry seedlings.</jats:p>