Mammalian cell lines can be efficiently established in vitro upon expression of the SV40 large T antigen driven by a promoter sequence derived from the human vimentin gene

Abstract The aim of this study was to investigate a new method to enhance the efficiency to create mammalian cell lines. Cell immortalization was achieved by intranuclear microinjection of a recombinant DNA construct composed of a constitutive promoter controlling the genes encoding immortalizing proteins; the sequences coding for the large T and small t antigens were fused downstream of regulatory elements from the vimentin gene, the activation of which characterizes the vast majority of cells growing in vitro . Data show that the efficiency of the immortalizing procedures using the SV40 early genes could be enhanced by the control elements derived from the human vimentin (HuVim) 5′ sequences that contained nucleotides −878 to +93 from the CAP site. This HuVim 830−T/t recombinant was used to create cell lines from numerous primary cultures of different origins: rabbit, porcine and human endothelial cells, rabbit and bovine epithelial cells. A set of large T-expressing cells was derived, and these cells retained characteristics of differentiated cells: binding of Ulex europaeus lectin and synthesis of Factor VIII for human endothelial cells; network of cytokeratin for bovine oviductal cells and rabbit mammary cells.