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Adenovirus vector designed for expression of toxic proteins
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
2001 (English)In: Journal of Virology, ISSN 0022-538X, E-ISSN 1098-5514, Vol. 75, no 20, 9579-9584 p.Article in journal (Refereed) Published
Abstract [en]

To construct recombinant adenoviruses expressing biologically active proteins may be impossible, or result in a significant reduction in virus yield, if the protein expressed has an inhibitory effect on virus replication or cellular growth. To overcome this problem, we previously designed adenovirus vectors expressing foreign proteins from inducible promoters. However, during our work with a replication-deficient virus expressing the ASF/SF2 splicing factor from a progesterone antagonist-inducible gene cassette, we discovered that ASF/SF2 was expressed at a significant level in the 293 producer cell line, even in the absence of inducer. 293 cells code for adenovirus E1A and E1B proteins and thus support the growth of E1-deficient adenoviruses. Here we show that this background ASF/SF2 expression results from a low level of E1A-mediated transactivation of the basal promoter driving transgene expression. To overcome the problem of leaky expression, we reconstructed a novel gene cassette that combines an inducible promoter and a Lac repressor protein-based block to reduce transcriptional elongation. We show that this novel vector system dramatically reduced background transgene expression and therefore should be useful for the rescue and propagation of high-titer stocks of recombinant adenoviruses expressing toxic proteins.

Place, publisher, year, edition, pages
2001. Vol. 75, no 20, 9579-9584 p.
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-52468DOI: 10.1128/JVI.75.20.9579-9584.2001ISI: 000171102900003PubMedID: 11559789OAI: oai:DiVA.org:uu-52468DiVA: diva2:80377
Available from: 2008-10-17 Created: 2008-10-17 Last updated: 2017-12-04Bibliographically approved

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Akusjärvi, Göran

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