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Flexible pseudotyping of retrovirus using recombinase-mediated cassette exchange
- Soares, HugoR., Almeida, AnaI., Tomás, HélioA., Alves, PaulaM., Coroadinha, AnaS.
- Biotechnology letters 2018 v.40 no.4 pp. 633-639
- Gammaretrovirus, cell lines, gene expression, proteins, research and development, transgenes
- OBJECTIVE: Develop an engineered cell line containing two flexible gene expression systems enabling the continuous production of tailor-made recombinant gammaretrovirus with predictable productivities through targeted integration. RESULTS: Dual-FLEX cells (dFLEX) contain two independent recombinase-mediated cassette exchange (RMCE) systems which confer flexibility to the expression of different transgene and envelope combinations. The flexible envelope expression in dFLEX cells was validated by pseudotyping retrovirus particles with three different viral envelope proteins—GaLV, 4070A and VSV-G. Our results show that dFLEX cells are able to provide high titers of infectious retroviral particles with a single-copy integration of the envelope constructs after RMCE. The integrated CRE/Lox tagging cassette was amenable to express envelope proteins both using constitutive (i.e. CMV) and inducible (i.e. Tet-on) promoters. CONCLUSIONS: dFLEX cell line provides predictable productivities of recombinant retrovirus pseudotyped with different envelope proteins broadening the tropism of particles that can be generated and thus accelerating the research and development of retrovirus-based products.