A controlled rather than an unreliable spontaneous in vitro flowering system of Gentiana triflora was established successfully. In vitro flower of gentian is of great similarity to in vivo flower, lending support to the former as an alternative in the proposed study on gentian flower physiology. The flowers can be obtained from more than 80% of the in vitro-cultured nodal explants in 12 weeks. Pulsing with 87.6 mM sucrose or 0.5 mM STS solution clearly improved bud opening and flower vase life in cut gentian. Total soluble sugar concentration increased markedly and starch content was obviously low in the petals after the sucrose treatment and this was confirmed using radioactive 14C-sucrose pulsing treatment. Exogenous ethylene could induce autocatalytic ethylene production in gentian petal and resulted in shortened flower vase life, while ethylene production of the flowers could be suppressed by the sucrose or STS treatment. Amylase activities were signifIcantly higher in 3% sucrose (w/v) or 0.5 mM STS treated flowers than in water treated control samples. Gentian flower senescence was associated with reduction of the protein content in the petals. Sucrose or STS pulsing treatment could increase protein content in the petals in comparison to that of water treated sample. A new polypeptide of low molecular weight was detected on SDS-PAGE gels used for analysis of the protein profIles of the petals treated with sucrose or STS. This suggested the possible changes in gene expression resulting from the pulsing treatments. Finally, a 931-bp cDNA fragment was obtained using a pair of primers designed from the conserved region of ACC oxidase gene sequences from several flowers using RT-PCR technique. Dot blot data suggested that ACC oxidase gene expression in the petal of cut gentian flowers could be suppressed by the administration of sucrose or STS. This indicated that changes in petal physiology in response to sucrose or STS were associated with ethylene-induced alterations in gene expression. Both sucrose and STS could be inhibitors of ethylene biosynthesis via regulating ACC oxidase gene expression.