General principles of retrovirus vector design.

An understanding in the life cycle of γ-retroviruses has led to significant progress in the development of murine leukemia virus (MLV)-based vectors for gene delivery and human gene therapy. An MLV-based vector consists of the cis-acting sequences important for viral replication and gene expression. The sequence that encodes viral proteins is replaced with the gene of interest. To generate infectious retroviral vectors, viral-encoded proteins are supplied in trans for virion assembly. Here, we describe a method to rapidly generate MLV vectors from transiently transfected human 293T cells. The strategies to purify and titer the vector and to detect the presence of replication competent retrovirus (RCR) in the vector harvest are also described. Copyright Â