The formation of flowers has been studied in stem tissue excised from flowering plants of Nicotiana tabacum variety Wisconsin No. 38, and cultured in vitro on Murashige and Skoog nutrient medium. A procedure for quantitative evaluation of factors influencing floral expression has been developed and effects of the growth substances, indole-3-acetic acid (IAA), kinetin and gibberellic acid (GA(3)), on the process are reported.Although a low (1 mum) level of IAA was required for the development of normal flowers on stem segments, higher concentrations tended to inhibit flowering. The decrease in floral buds was rapid in the 3 to 15 mum range. IAA concentrations up to 75 mum increased vegetative bud formation so as to effect a transition from floral to vegetative buds rather than merely an inhibition of bud formation. Higher IAA concentrations inhibited both vegetative and floral bud formation.Kinetin in high concentrations greatly increased the number of vegetative buds but had no significant effect on the number of floral buds per segment. High kinetin concentrations also permitted branching of floral shoots so that flower clusters were formed.GA(3) applied in the medium from the start, strongly inhibited bud formation on the stem segments, but when applied to young floral buds after they had formed, it promoted their further development ("bolting").Flower formation occurred in complete darkness, but light of moderate intensity was required for the development of normal flowers.