Show simple item record

dc.contributor.authorSchucht, R
dc.contributor.authorCoroadinha, A S
dc.contributor.authorZanta-Boussif, M A
dc.contributor.authorVerhoeyen, E
dc.contributor.authorCarrondo, M J T
dc.contributor.authorHauser, Hansjoerg
dc.contributor.authorWirth, Dagmar
dc.date.accessioned2007-11-16T14:34:49Z
dc.date.available2007-11-16T14:34:49Z
dc.date.issued2006-08-01
dc.identifier.citationMol. Ther. 2006, 14(2):285-92en
dc.identifier.issn1525-0016
dc.identifier.pmid16697259
dc.identifier.doi10.1016/j.ymthe.2005.12.003
dc.identifier.urihttp://hdl.handle.net/10033/14594
dc.description.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.
dc.format.extent387713 bytes
dc.format.mimetypeapplication/pdf
dc.language.isoenen
dc.titleA new generation of retroviral producer cells: predictable and stable virus production by Flp-mediated site-specific integration of retroviral vectors.en
dc.typeArticleen
dc.format.digYES
refterms.dateFOA2018-06-12T21:40:14Z
html.description.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.


Files in this item

Thumbnail
Name:
Publisher version
Thumbnail
Name:
Schucht_final.pdf
Size:
378.6Kb
Format:
PDF
Description:
original document

This item appears in the following Collection(s)

Show simple item record