Regenerating calli of Allium fistulosum × A. cepa interspecific F1 hybrids were treated in vitro with colchicine. A factorial experiment (colchicine concentration × time) was used to recover tetraploids from calli treated with colchicine in vitro. Shoot production of regenerating calli following in vitro colchicine treatment decreased with increasing colchicine concentration and treatment time. Cytological analyses of root tip cells from regenerated plantlets showed that chromosomes of control plantlets (not treated with colchicine) were not doubled. Chromosomes of some plantlets regenerated from in vitro colchicine treated calli were doubled, resulting in tetraploids. Calli treated with 0.1 or 0.2% colchicine in BDS (Dunstan & Short, 1977) liquid medium for 48 or 72 hours yielded the highest number of tetraploid plantlets. These results demonstrate that in vitro colchicine treatment of regenerating calli of interspecific F1 hybrids is effective in recovering tetraploids.