A rapid clonal propagation method was developed for Centaurea ragusina L. (Asteraceae), a rare Croatian endemic species. Shoots originated from aseptically germinated seeds were used for culture initiation. The highest multiplication rate, 4.6 shoots per explant, was achieved in a 4-week culture period on half-strength Murashige and Skoog medium supplemented with 1.0 µM 6-benzylaminopurine and 2.9 µM gibberellic acid. Excised shoots were rooted on the same basal medium with 2.5 µM indole-3-butyric acid added. Rooted plantlets were successfully transferred to potting soil and acclimatized to outdoor conditions.