Recent Developments in Retroviral-Mediated Gene Transduction Gordon Daly, Yuti Chernajovsky  Molecular Therapy  Volume 2, Issue 5, Pages 423-434 (November 2000) DOI: 10.1006/mthe.2000.0211 Copyright © 2000 American Society for Gene Therapy Terms and Conditions

FIG. 1 Packaging of replication deficient retroviral vector in packaging cells. Packaging cells engineered to express retroviral structural genes devoid of packaging signal (ψ) are transfected with plasmids containing the retroviral vector which includes the ψ sequence. Upon transcription of the retroviral vector mRNA it gets packaged into viral particles which bud from the cell surface and can infect target cells. Molecular Therapy 2000 2, 423-434DOI: (10.1006/mthe.2000.0211) Copyright © 2000 American Society for Gene Therapy Terms and Conditions

FIG. 2 Basic structure of the plasmoviral vector system. The retroviral vector is flanked by LTRs and includes the ψ-packaging signal, Gag/Pol genes and a reporter gene or other gene of interest. 3’ to the vector, the plasmid contains a CMV transcriptional unit driving expression of amphotropic Env. As this Env is not included between the LTRs, the retrovirus packaged can not be transmitted to additional cells once incorporated into its primary target cell. Molecular Therapy 2000 2, 423-434DOI: (10.1006/mthe.2000.0211) Copyright © 2000 American Society for Gene Therapy Terms and Conditions

FIG. 3 Genome organization of HIV. Between the LTRs of HIV a complex genome expressing Gag, Pro/Pol, Vpr (R), Vif, Vpu (U), Tat, Rev, and Nef is assembled. Several splice donor and acceptor sites (shown as separating black flat surfaces) ensure the expression of several mRNAs capable of translating these gene products. The functions of these genes are discussed in the text. Molecular Therapy 2000 2, 423-434DOI: (10.1006/mthe.2000.0211) Copyright © 2000 American Society for Gene Therapy Terms and Conditions

FIG. 4 Schematic representation of the first-generation lentiviral vector system developed by Naldini et al. (107). The three plasmid system consists of two packaging plasmids. One expresses either the VSV-G protein or the amphotropic Env gene, the second provides all HIV proteins excluding Env and Vpu and cannot encapsidate as has its ψ element deleted. The third plasmid is the lentiviral transfer vector per se containing between the LTRs the ψ sequence with a truncated Gag gene out of frame, an RRE element for efficient nuclear transport and an internal CMV promoter for expression of a gene of interest. Molecular Therapy 2000 2, 423-434DOI: (10.1006/mthe.2000.0211) Copyright © 2000 American Society for Gene Therapy Terms and Conditions