Immortalization by gene transfection.

Cultured cell lines that maintain specific differentiated phenotypes have been indispensable tools in cell biology. Progress in understanding the function of differentiated cells in vivo can be facilitated by creating cell lines via immortalizing gene transduction, if they retain the essential differentiated features of the same cells in vivo. Rodent cells immortalize spontaneously with a frequency of 10(-5) to 10(-6). Thus, it is easy to isolate immortal cells from rodent cell populations even without the transfer of immortalizing genes. Immortalizing genes can be used to increase this frequency to approximately 100%. In contrast, the spontaneous immortalization of human cells is a very rare event; the frequency is thought to be < 10(-12). Immortalizing genes can also be used to increase this frequency. Several genes that promise efficient immortalization of cultured cells have been identified. Immortalizing genes include simian virus 40 large T antigen, papillomaviruses E6 and E7, adenovirus E1A, Epstein-Barr virus, human T-cell leukemia virus, herpesvirus saimiri, oncogenes, and mutant p53 gene. Equally important, innovative means of gene delivery have been developed as well. These immortalizing genes, together with gene transfer methodologies, have provided the means to generate cell lines from cell types that are not abundant or are difficult to obtain in pure form in primary culture, are in short supply as human cells, and/or have brief lifetimes in culture. This chapter focuses primarily on the immortalization method by gene transfection. The chapter is not meant to be comprehensive, but rather to provide an account of the power and usefulness of immortalization methodology.