Financial support: National Institutes of Health, USA.

Keywords: 10T1/2 cells, Ras, Tetracycline-inducible expression system, tetR, T-REx.

Abbreviations:
PCMV: cytomegalovirus enhancer-promoter;
tetO: tetracycline operator sites;
TetR: tetracycline repressor.

The technology of tetracycline-inducible gene expression has been successfully used in experimental biology to identify the function and downstream signaling pathways of an interested gene. It has been significantly improved to meet the criteria with specificity to exogenous non-toxic inducers, independent regulation from cellular pathways, and dose-dependent inducibility and reversibility. However, to establish tetracycline-inducible gene expression in mammalian cells is still a time-and effort-consuming process. With a tetracycline-inducible gene expression system T-REx, we have developed a practical protocol to use the oncogenic H-ras gene as a dominant reporter gene to increase efficiency in attaining desired cell lines in which ectopic expression of a particular gene in cells can be introduced and reversibly induced by the presence or absence of tetracycline in cultures.