Ficus carica L. (Fig) is one of the earliest cultivated fruit trees and is extensively grown in the Mediterranean, Middle East, and Egypt's desert regions. Despite its importance, Fig cultivation faces challenges, including low propagation success rates and Fig Mosaic Virus (FMV) infections. This study aimed to overcome these hurdles through biotechnological approaches and the use of Artemisia annua extract. Virus-free Fig plants were generated from infected 'Sultani' cultivars using tissue culture techniques. Additionally, the antiviral properties of A. annua extract against FMV were assessed. The extract markedly reduced oxidative stress biomarkers, such as MDA, carbonyl, and H2O2, and decreased the expression levels of defence-related genes, including PAL, PPO, PPX, PR3, PR5 and Tubulin, in a dose-dependent manner, with concentrations of 200 and 400 mu l/l showing the most significant effects. Results indicated a reduction of 40-60% in oxidative stress markers and a general reduction in gene expression fold changes in virus-infected samples treated with the extract compared to controls. These findings highlight the potential of integrating biotechnological techniques and plant-derived compounds to enhance Fig cultivation, ensuring sustainable production and improved plant health. The successful propagation of virus-free Fig plants and the efficacy of A. annua extract in combating FMV showcase promising methodologies to address current propagation difficulties and viral threats in Fig agriculture. This research underscores the importance of biotechnological innovations and the use of natural antiviral agents in advancing agricultural practices for global economic benefits.