SpringerProtocols: Abstract: Cytogenetic Characterization of Recombinant Cells

Cytogenetic Characterization of Recombinant Cells

By: Florian M. Wurm², Dietmar Schiffmann²

Abstract

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The use of mammalian cell systems for recombinant protein production has become the dominant method to produce high-value proteins for therapeutic purposes. Recombinant cell lines are often generated by transfection with specific DNA; therefore a selectable system is desirable to discriminate cells that have integrated the foreign DNA. Chinese hamster ovary (CHO) cells are most commonly used because of the availability of both a dihydrofolate reductase (DHFR)-deficient mutant cell line (CHO-DUKX (1) and DHFR expression vectors. Since DHFR expression vectors can be linked to DNA coding for desired proteins, co-integration of the DHFR expression vectors and the associated DNA often occurs. Integration of the DHFR expression vector into the CHO genome enables the cells to grow in a medium lacking the components glycine, hypoxanthine, and thymidine. Also, when multiple copies of the DHFR expression vector have been integrated, cellular resistance to methotrexate (MTX), a folate analog, is mediated. MTX can therefore be used to select for subclones that have amplified the transfected plasmid sequences to very high copy numbers.

Affiliation(s): (2) Institut de Genie Chimique, Ecole Polytechnique Federale de Lausanne, Lausanne, Switzerland

Book Title: Animal Cell Biotechnology: Methods and Protocols

Series: Methods in Biotechnology | Volume: 8 | Pub. Date: Feb-22-1999 | Page Range: 49-59 | DOI: 10.1385/0-89603-547-6:49

Subject: Biotechnology

References

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