Amplification using CHO cell expression vectors.
The ability to select for integration of plasmid DNA into the host chromosome allows the generation of stably transfected cell lines. With transfection of a selectable marker linked to a nonselectable target gene (or by cotransfection of the two unlinked genes), high-level expression of the desired gene is obtained by selecting for amplification of the selectable marker. This unit presents two systems for gene amplification and expression. The first describes the dihydrofolate reductase (DHFR) selection system while the second is based on selection of the glutamine synthetase (GS) gene. The DHFR system is probably more widely used, and results in very high levels of amplification and expression; however, the DHFR amplification process is lengthy and may require several months to isolate and characterize a stable, amplified line. In contrast, the GS system typically requires only a single round of selection for amplification to achieve maximal expression levels.
Kingston
Robert E
RE
Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts, USA.
Kaufman
Randal J
RJ
Bebbington
C R
CR
Rolfe
M R
MR
D000818:Animals; D016466:CHO Cells / Q000187:drug effects / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D005784:Gene Amplification; D005822:Genetic Vectors / Q000235:genetics*; D005974:Glutamate-Ammonia Ligase / Q000235:genetics; D007202:Indicators and Reagents; D008727:Methotrexate / Q000494:pharmacology; D012641:Selection, Genetic; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D014162:Transfection / Q000379:methods; D016662:Virus Integration
D007202:Indicators and Reagents; D013762:Tetrahydrofolate Dehydrogenase; D005974:Glutamate-Ammonia Ligase; D008727:Methotrexate