Figure 1.

Schematic representation of nonintegrated lentiviral (NILV) circles in which 2-LTRs have joined by nonhomologous end-joining in the absence of integrase activity. These episomal forms support transcription and one attractive application has been the delivery of nucleases capable of site-specific DNA modification. In the case of zinc finger nucleases (ZFNs), the zinc finger motifs bind to opposite strands of target DNA sites and this allows dimerization of Fok1 endonuclease resulting in double-stranded cleavage of genomic DNA. Alternatively, meganucleases (MN) mediate precise DNA cleavage following highly specific DNA recognition and binding. Provision of a third NILV, carrying sequences homologous to the cleavage site, provides a template for homologous recombination and repair of cleaved genomic DNA, and this can allow the insertion of a minimal promoter/transgene cassette if flanked by appropriate homology sequences.