Oncoretroviral vectors and lentiviral vectors offer the potential for long-term gene expression by virtue of their stable chromosomal integration and lack of viral gene expression. Consequently, their integration allows passage of the transgene to all progeny cells, which makes them particularly suitable for stem cell transduction. However, a disadvantage of oncoretroviral vectors based on Moloney murine leukemia virus (MoMLV) is that cell division is required for transduction and integration, thereby limiting oncoretroviral-mediated gene therapy to actively dividing target cells. In contrast, lentiviral vectors can transduce both dividing and nondividing cells. Lentiviral vectors have been derived from either human or primate lentiviruses, with the human immunodeficiency virus (HIV) as prototype, or from nonprimate lentiviruses, such as the equine infectious anemia virus (EIAV). The ability to pseudotype oncoretroviral and lentiviral vectors with the vesicular stomatitis virus G glycoprotein (VSV-G) allowed for the production of high-titer vectors (10(9)-10(10) transducing units/ml). These high-titer vector preparations were shown to effectively cure genetic diseases in experimental animal models and constitute an essential step toward direct in vivo gene therapy applications. This chapter focuses on different methods that permit large-scale production of high-titer VSV-G pseudotyped oncoretroviral and primate or nonprimate lentiviral vectors and highlights their importance for achieving therapeutic effects in preclinical animal models.