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Gewählte Publikation:

Zboray, K; Sommeregger, W; Bogner, E; Gili, A; Sterovsky, T; Fauland, K; Grabner, B; Stiedl, P; Moll, HP; Bauer, A; Kunert, R; Casanova, E; .
(2015): Heterologous protein production using euchromatin-containing expression vectors in mammalian cells.
Nucleic Acids Res. 2015; 43(16):e102 FullText FullText_BOKU

Upon stable cell line generation, chromosomal integration site of the vector DNA has a major impact on transgene expression. Here we apply an active gene environment, rather than specified genetic elements, in expression vectors used for random integration. We generated a set of Bacterial Artificial Chromosome (BAC) vectors with different open chromatin regions, promoters and gene regulatory elements and tested their impact on recombinant protein expression in CHO cells. We identified the Rosa26 BAC as the most efficient vector backbone showing a ninefold increase in both polyclonal and clonal production of the human IgG-Fc. Clonal protein production was directly proportional to integrated vector copy numbers and remained stable during 10 weeks without selection pressure. Finally, we demonstrated the advantages of BAC-based vectors by producing two additional proteins, HIV-1 glycoprotein CN54gp140 and HIV-1 neutralizing PG9 antibody, in bioreactors and shake flasks reaching a production yield of 1 g/l.
Autor/innen der BOKU Wien:
Kunert Renate
Sommeregger Wolfgang
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