AN EPISOMAL VECTOR FOR STABLE TETRACYCLINE-REGULATED GENE-EXPRESSION

The recently introduced tetracycline (Tc)-regulatable eukaryotic gene expression system based on the Escherichia coil Tn 10 tetracycline ope ron has proven to be a powerful tool for controlled expression of a va riety of genes in vitro as well as in vivo, Control elements of this e xpression system are contained in two separate plasmid vectors, The tT A vector encodes a transactivator protein and the tetP vector contains a responsive operator-promoter element (tetP) that controls gene expr ession depending on tTA binding, Establishment of cell lines expressin g a gene of interest under tetP control requires two subsequent rounds of transfection and clonal selection after each transfection. Here we describe a modification of this system in which the tetP element is p laced in an episomal EBNA-based plasmid that can be stably maintained in primate but not in rodent cells, Using HeLa and human melanoma cell s, we show that upon transient or stable transfection a reporter gene is expressed in a Tc-regulated manner similar to the original system, Thus, this expression system combines the advantages of episomal vecto rs, such as high efficiency of transfection and time-efficient selecti on of mass cultures, with tight control of gene expression provided by the Tc-regulatable system.