A novel family of retroviral vectors for the rapid production of complex stable cell lines

The production of stable cell lines is an important technique in cell biolo gy, and it is often the rate-limiting step in studies involving the charact erization of the function of novel genes or gene mutations. To facilitate t his process, a novel family of retroviral vectors, the pE vector family, ha s been generated. The retroviral sequences in the pE vectors have been take n from the Moloney murine leukemia virus (MMLV) vector pMFG, which has been shown to express cDNA inserts more consistently and at higher levels than earlier generations of MMLV vectors. These vectors contain four different i nternal ribosome entry site-selectable markers, allowing high-efficiency se lection of transductants expressing the desired cDNA. The pE vectors have a n episomal design to allow long-term production of high-titer virus without the need for subcloning the producer line. Using a strategy of combinatori al infection followed by combinatorial drug selection, we demonstrate that the pE vectors can be used to generate stable, polyclonal cell lines expres sing at least three novel cDNAs in less than 2 weeks. The use of these vect ors will thus dramatically accelerate the production of complex stable cell lines. (C) 2001 Academic Press.