Ploidy stability in gynogenic clones of long-day onion is assessed during micropropagation. In order to induce chromosome doubling in haploids, explants and plantlets of three clones were treated in vitro. Colchicine (0.625-12.5 mM) and oryzalin (10-200 μM) were tested for 24 h in combination with culture conditions (4°C in the dark or 24°C in the 16 h day photoperiod). Diploidization performed well (64%) during micropropagation on explants treated with 0.625 mM colchicine at 24°C day. Among treatments on individual plantlets, best results were obtained either with 2.5 mM colchicine (up to 65.7% of diploids) or 50 μM oryzalin (up to 57.1% of diploids). Both chemicals induced mixoploids and affected plant regeneration, but a better plant quality was obtained with oryzalin. Chromosome doubling in explants during the first cycle of micropropagation after gynogenesis appears to be a valuable and time-saving procedure to provide numerous doubled-haploid plants.