Figure 4.

System for Inducible Expression of Epitope-Tagged ACS5.

(A) Diagram of the inducible system used to express ACS5. The DEX-inducible system (Aoyama and Chua, 1997) uses a tripartite fusion transcription factor (GVG fusion) composed of the GAL4 DNA binding domain, the VP16 activation domain, and the glucocorticoid-responsive domain to drive DEX-inducible expression of the target gene. We placed the ACS5 coding region fused to a 6 × myc epitope tag downstream of the GAL4 operator sequences to which the GVG fusion protein binds, which results in DEX-responsive expression of the fusion protein.

(B) Phenotypes of 3-day-old etiolated seedlings from a transgenic line harboring the myc-ACS5^WT transgene grown in the presence (+) and absence (−) of DEX. Seedlings were grown on MS medium, and representative seedlings were photographed.

(C) Measurement of ethylene produced by 3-day-old etiolated seedlings of a myc-ACS5^WT transgenic line grown on MS medium in the presence of increasing concentrations of DEX. The inset shows an immunoblot using the anti-myc monoclonal antibody as a probe of protein extracts from 3-day-old etiolated seedlings of the same line grown at the indicated concentrations of DEX.