Simian retrovirus vectors for gene transfer in nonhuman primate cells.
We have recently identified and sequenced a molecular clone of the serogroup 2 simian retrovirus (SRV), D2/RHE/OR/V1, that retains an enhanced ability to infect specific T cell lines. In this report, using deletion mutagenesis, we localized the psi packaging signal, necessary for packaging of D2/RHE/OR/V1 particles, to the genomic region 345-650, which comprises the 5' intergenic region (IR) and the extreme 5' portion of the gag gene. To build an SRV-based gene transfer system and to reduce the possibility of recombination and regeneration of replication-competent viruses, we constructed split-genome D2/RHE/OR/V1 plasmid recombinants containing distinct and non-overlapping retroviral gene regions and several replacement components. For the retrovirus gene transfer vehicle, we deleted the D2/RHE/OR/V1 structural genes and substituted a cassette including the psi-packaging region, the beta-galactosidase reporter gene, and the 3' IR. Both packaging cell recombinants were used to generate stable monkey packaging cell lines; the gene transfer vehicle was subsequently transfected into the packaging cell lines, and replication-defective viruses were recovered for subsequent infection into fresh monkey cells. Successful infection by the recovered viruses verifies the potential efficacy of the SRV-based system as a research tool for gene transfer of heterologous genes into nonhuman primate cells.
Li
B
B
Division of Neuroscience, Oregon Regional Primate Research Center, Oregon Health Sciences University West Campus, 505 NW 185th Avenue, Beaverton, OR 97006, USA.
Nguyen
S
S
Li
X
X
Machida
C A
CA
D020121:5' Untranslated Regions / Q000235:genetics; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002999:Clone Cells; D004274:DNA, Recombinant / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D015683:Gene Products, gag / Q000235:genetics; D018014:Gene Transfer Techniques; D017930:Genes, Reporter / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics*; D016679:Genome, Viral; D006801:Humans; D007763:Lac Operon / Q000235:genetics; D051381:Rats; D015301:Retroviruses, Simian / Q000235:genetics*; D014161:Transduction, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D014771:Virion / Q000235:genetics / Q000378:metabolism; D019065:Virus Assembly / Q000235:genetics; D001616:beta-Galactosidase / Q000235:genetics
D020121:5' Untranslated Regions; D004274:DNA, Recombinant; D004279:DNA, Viral; D015683:Gene Products, gag; D001616:beta-Galactosidase