Generation of human cell lines using lentiviral-mediated genetic engineering.

Even now, most human cell lines used in research are derived from tumor cells. They are still widely used because they grow well in vitro and so far have helped answering several basic biological questions. However, as modern biology moves into more sophisticated areas, scientists now need human cell lines closer to normal primary cells and further from transformed cancerous cells. The recent identification of cellular genes involved in cell cycling and senescence, together with the development of molecular tools capable of cleanly integrating transgenes into the genome of target cells, have moved the frontier of genetic engineering. In this chapter, we present a detailed hands-on protocol, based on lentivirus-derived vectors and a combination of two native cellular genes that has proven very efficient in generating immortal cell lines from several human primary cells, while preserving most of their original properties.