The Glucocorticoid Inducible Expression Of The Green Fluorescent Protein (GFP) In The Tobacco

The glucocorticoid inducible expression system is based on the chimerical transcription factor, designated as GVG, which consists of the DNA-binding domain of the yeast transcription factor GAL4, the transactivating domain of the herpes viral protein VP16, and the receptor domain of the rat glucocorticoid receptor (GR). In the absence of its ligand, glucocorticoid hormone, GVG is produced and resides in the cytoplasm, where it forms complexes with multiple proteins. After binding to glucocorticoid, the modified GVG protein is released from the complexes, transferred into the nucleus and regulates specific gene expression. The binary vector pIndex3, carrying the glucocorticoid inducible expression system, is transferred into Agrobacterium in order to transform wild-type tobacco. After the detection, it is sure that the glucocorticoid inducible expression system has been established. In this system, the transcription factor GVG gene under the control of the 35S promoter and the mgfp5 transcribed from an inducible promoter containing four tandem copies of the GAL4 upstream activating sequence and a truncated 35S promoter. As the target gene of the chemically inducible system, the GFP expression is stringently controlled. In the presence of the inducer, the mRNA of mgfp5 was transcribed and the GFP began to express around the veins where the inducer (dexamethasone, Dex, a kind of man-made glucocorticoid) reached early. When the inducer was discarded, the GFP expression was stopped. According to the experiment, the young leaves were more sensitive to the inducer than the old ones. To the next experimental step, the non-translated region of the PVX was inserted flanking the mgfp5 gene and it did not repress the GFP expression. The GVG system has the merits, high stringency and specificity, and controls the transgene expression temporally, spatially and quantitatively with the help of exogenous chemicals. The establishment of this system offers the possibility for both theoretical and practical applications by the chemically inducible system.