Lentiviral Vectors Preparation and Use

Lentiviruses, such as human immunodeficiency virus (HIV), feline immunodeficiency virus (FIV), and equine infectious anemia virus (EIAV), are members of the Retroviridae, viruses with enveloped capsids and a plus-stranded RNA genome. Like all retroviruses, the RNA genome of lentivirus is converted to DNA by reverse transcription after infection and is subsequently stably integrated into the host cell genome. However, unlike other retroviruses, which mainly infect dividing cells, lentiviruses can infect both dividing and nondividing cells (1 –3 ). Gene therapy viral vectors are produced by cotransfection of plasmids encoding viral envelopes, capsids, enzymes, and viral RNA. The specific tissue tropism of the lentiviral envelope restricts its use as a gene transfer vector. This limitation has been overcome by pseudotyping the viral envelope with vesicular stomatitis virus G (VSV-G) glycoprotein (4 ,5 ). The aim of this chapter is to describe the general procedures developed for the preparation and use of an HIV type 1 (HIV-1)-based lentiviral vector system. Detailed descriptions of lentiviral molecular biology can be found in several recent reviews (1 ,6 ,7 ).