P96: Sensitivity Enhancement of Fluorescent Protein Based reporter gene system by Cyclin B1 Destruction Box

The regulation of genes is often difficulty observed in cells. Therefore, to overcome such problem, the report gene system was developed to examine the precise gene regulations in cells and the system also can be used as functional compound screening tools. Fluorescence proteins are frequently used in report gene system, because of its convenience feature with simplicity, rapidity and clarity. However, fluorescence proteins are overly stable that cause its insensitivity of analysis. Besides, luciferase and secreted alkaline phosphatase (SEAP) are frequently used enzymes in report gene system. Although luciferase is a sensitive enzyme for analysis of gene regulation, but the analytic procedures are complex and time-consuming. Cyclin destruction box (CDB) is a protein fragment of cyclin in cells, which leads cyclin degradation under the control of cell cycles. Thus, CDB has the function to decrease the stability of its target fusion proteins. In this study, we use the CDB to modify the common used report gene system to establish novel assay system with fluorescence proteins. Our results demonstrated that both green and red based fluorescence protein will decrease its stability in the transfected cells. Moreover, the gene regulative sensitivity of such CDB modified system was enhanced when the promoter inducer was added to cells. We supposed that the improved gene reporter system can be utilized as a tool for functional compound screening in the future.