Regulation of gene expression in mammalian cells using the lac repressor

摘要

The study describes the construction of a one step inducible lac repressor/operator mammalian expression system within the context of DNA mediated vaccination (naked DNA). It is novel in that it contains, on a single vector (pSO1), all the cis and trans controlling elements necessary to manipulate (switch on/off) and control the expression of a reporter gene in a mammalian cell. The enhanced green fluorescent protein (EGFP) gene was cloned downstream of the cytomegalovirus immediate-early enhancer promoter (PCMV IE) and a lac operator (LacO), which was in turn regulated by the expression of the lac repressor under the control of a second PCMV IE. The number of cells fluorescing were greatly increased following isopropy B-D thiogalactoside (IPTG) induction. In the repressed state, fewer pSO1 transfected cells expressed the EGFP, and the fluorescence intensity was also lower than that observed for the induced pSO1 transfected cells. Observation by microscopy was quantified by FACScan analysis on the different populations of cells.