A new generation of retroviral producer cells: predictable and stable virus production by Flp-mediated site-specific integration of retroviral vectors.
Cast your vote
You can rate an item by clicking the amount of stars they wish to award to this item.
When enough users have cast their vote on this item, the average rating will also be shown.
Your vote was cast
Thank you for your feedback
Thank you for your feedback
Coroadinha, A S
Zanta-Boussif, M A
Carrondo, M J T
MetadataShow full item record
AbstractWe developed a new strategy that provides well-defined high-titer producer cells for recombinant retroviruses in a minimum amount of time. The strategy involves the targeted integration of the retroviral vector into a chromosomal locus with favorable properties. For proof of concept we established a novel HEK293-based retroviral producer cell line, called Flp293A, with a single-copy retroviral vector integrated at a selected chromosomal locus. The vector was flanked by noninteracting Flp-recombinase recognition sites and was exchanged for different retroviral vectors via Flp-mediated cassette exchange. All analyzed cell clones showed correct integration and identical titers for each of the vectors, confirming that the expression characteristics from the parental cell were preserved. Titers up to 2.5 x 10(7) infectious particles/10(6) cells were obtained. Also, high-titer producer cells for a therapeutic vector that encodes the 8.9-kb collagen VII cDNA in a marker-free cassette were obtained within 3 weeks without screening. Thus, we provide evidence that the precise integration of viral vectors into a favorable chromosomal locus leads to high and predictable virus production. This method is compatible with other retroviral vectors, including self-inactivating vectors and marker-free vectors. Further, it provides a tool for evaluation of different retroviral vector designs.
CitationMol. Ther. 2006, 14(2):285-92
- The use of recombinase mediated cassette exchange in retroviral vector producer cell lines: predictability and efficiency by transgene exchange.
- Authors: Coroadinha AS, Schucht R, Gama-Norton L, Wirth D, Hauser H, Carrondo MJ
- Issue date: 2006 Jul 13
- An improved method for generating retroviral producer clones for vectors lacking a selectable marker gene.
- Authors: Persons DA, Mehaffey MG, Kaleko M, Nienhuis AW, Vanin EF
- Issue date: 1998 Jun
- Retargeting of retroviral integration sites for the predictable expression of transgenes and the analysis of cis-acting sequences.
- Authors: Schübeler D, Maass K, Bode J
- Issue date: 1998 Aug 25
- Site-specific integration of targeted DNA into animal cell genomes.
- Authors: Koch KS, Aoki T, Wang Y, Atkinson AE, Gleiberman AS, Glebov OK, Leffert HL
- Issue date: 2000 May 16
- Flp-mediated integration of expression cassettes into FRT-tagged chromosomal loci in mammalian cells.
- Authors: Wirth D, Hauser H
- Issue date: 2004