Generation of High-Titer Pseudotyped Lentiviral Vectors

Lentiviral vectors (LVs) are widely used in gene transfer protocols due to many advantages that include stable gene expression, higher transgene payloads, and, importantly, the ability to pseudotype the vectors with a diverse number of heterologous viral envelopes with broad or restricted cell tropism depending on the need. The pseudotyping process also allows for incorporation of specific antibodies/ligands to engineer LVs. These features greatly facilitate customization of lentiviral vectors for cell/tissue specific gene delivery. The VSV-G protein containing envelope remains the most widely used among the viral glycoproteins used for LV pseudotyping due to its versatile host range and stability. However, many other viral envelopes are being identified for special applications of LVs. Here we describe the methodology to generate pseudotyped LVs using a four-plasmid transient transfection system focusing on aspects to generate high-titer vector stocks.