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. 2015 Apr 30;33(26):2955–2962. doi: 10.1016/j.vaccine.2015.04.071

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© 2015

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Fig. 3 — Development of the WinPac lentiviral vector packaging cell line. Transduction of a gammaretroviral vector into 293FT cells and integration of a hygro-eGFP gene into active genomic loci tagged by mutant LoxP sites. Isolation of a cell clone with a single, stable, highly-expressing genomic locus. Site-directed insertion of a codon-optimised HIV gag-pol construct using Cre-LoxP recombinase-mediated cassette exchange. and Stable transfection of plasmids encoding HIV rev, RDpro and a GFP-expressing, self-inactivating lentiviral vector (in tandem with pSELECT antibiotic resistance plasmid), prior to multi-antibiotic selection of cell clones actively expressing lentiviral vector constructs. LTR = long terminal repeat; Neo^R = neomycin resistance gene; CMV pro = cytomegalovirus promoter; Hygro-eGFP = hygromycin-resistant enhanced green fluorescent protein; Puro^R = puromycin resistance gene; SV40 polyA = simian virus 40 polyadenylation sequence; HIV gag-pol = human immunodeficiency virus gag-pol genes; Cre-RMCE = Cre-recombinase mediated cassette exchange; pTK = thymidine kinase promoter; Hygro^R = hygromycin resistance gene; HIV rev = human immunodeficiency virus rev gene; MLV = murine leukaemia virus; RDpro env = RD114-HIV chimeric virus envelope glycoprotein; Phleo^R = phleomycin resistance gene; Ψ = encapsidation signal; RRE = HIV rev response element; cPPT = HIV central polypurine tract; SFFV pro = spleen focus forming virus promoter; WPRE = Woodchuck hepatitis virus post-transcriptional regulatory element; BSr = blasticidin S-resistance gene; pA = polyadenylation sequence.

Inline graphic — Development of the WinPac lentiviral vector packaging cell line. Transduction of a gammaretroviral vector into 293FT cells and integration of a hygro-eGFP gene into active genomic loci tagged by mutant LoxP sites. Isolation of a cell clone with a single, stable, highly-expressing genomic locus. Site-directed insertion of a codon-optimised HIV gag-pol construct using Cre-LoxP recombinase-mediated cassette exchange. and Stable transfection of plasmids encoding HIV rev, RDpro and a GFP-expressing, self-inactivating lentiviral vector (in tandem with pSELECT antibiotic resistance plasmid), prior to multi-antibiotic selection of cell clones actively expressing lentiviral vector constructs. LTR = long terminal repeat; Neo^R = neomycin resistance gene; CMV pro = cytomegalovirus promoter; Hygro-eGFP = hygromycin-resistant enhanced green fluorescent protein; Puro^R = puromycin resistance gene; SV40 polyA = simian virus 40 polyadenylation sequence; HIV gag-pol = human immunodeficiency virus gag-pol genes; Cre-RMCE = Cre-recombinase mediated cassette exchange; pTK = thymidine kinase promoter; Hygro^R = hygromycin resistance gene; HIV rev = human immunodeficiency virus rev gene; MLV = murine leukaemia virus; RDpro env = RD114-HIV chimeric virus envelope glycoprotein; Phleo^R = phleomycin resistance gene; Ψ = encapsidation signal; RRE = HIV rev response element; cPPT = HIV central polypurine tract; SFFV pro = spleen focus forming virus promoter; WPRE = Woodchuck hepatitis virus post-transcriptional regulatory element; BSr = blasticidin S-resistance gene; pA = polyadenylation sequence.