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Utilization of an Epstein-Barr virus replicon as a eukaryotic expression vector.
J M Young1, C Cheadle, J S Foulke
1Molecular Biology Division, Rorer Biotechnology, Inc., Springfield, VA 22151.
Gene
|January 1, 1988
Summary
Epstein-Barr virus (EBV) replicons enable efficient foreign gene expression and protein secretion in mammalian cells. This system shows promise for biotechnological applications, particularly in human embryonic 293 cells.
Area of Science:
- Molecular Biology
- Virology
- Biotechnology
Background:
- Epstein-Barr virus (EBV) replicons, comprising oriP and EBNA-1, are maintained episomally in mammalian cells.
- EBV replicons offer a platform for stable episomal maintenance and gene expression.
Purpose of the Study:
- To evaluate the utility of an EBV replicon for foreign gene expression.
- To assess the efficiency of protein secretion using this system.
Main Methods:
- Cloning cDNAs for human interferon-gamma (IFN-gamma) and EGF receptor extracellular domain (EGF-Rex) into an EBV replicon vector.
- Expressing these constructs in human embryonic 293 cells and CV-1 cells.
- Measuring protein expression and secretion into extracellular media.
Main Results:
- Efficient expression and secretion of human IFN-gamma and EGF-Rex were achieved using the EBV replicon.
- Expression levels were approximately ten-fold higher in human embryonic 293 cells compared to CV-1 cells.
- Protein expression was dependent on the orientation of the IFN-gamma transcriptional cassette.
Conclusions:
- EBV replicons are effective tools for high-level foreign gene expression and protein secretion in mammalian cells.
- Human embryonic 293 cells provide a superior expression environment for this system.
- Vector design, specifically transcriptional cassette orientation, influences expression efficiency.