High efficiency transfection of monkey kidney COS-1 cells
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Methods are described for achieving high efficiency transient transfection of COS-1 cells. A transfection solution of vector DNA and DEAE-dextran in phosphate buffered saline is added to the cells, followed by treatment with culture medium containing chloroquine, resulting in a maximum efficiency of about 50%. The high efficiency obtained is primarily dependent on the use of Ca2+- or Mg2+-free buffer solutions for transfection, because the addition of these ions greatly reduces efficiency. Shocking the cells with dimethylsulfoxide does not increase transfection efficiency. COS-1 cells are monkey kidney cells that have a genomic insertion of the SV40 T antigen gene, allowing plasmid expression vectors bearing an SV40 replication origin to be amplified in these cells. Therefore this transfection method is useful for optimizing transient expression of genes in a mammalian cell system.
Key wordsDEAE-dextran transfection COS cells transient expression
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- 6.Hall, C. V.; Jacob, P. E.; Ringold, G. M., et al. Expression and regulation ofEscherichia coli lacZ gene fusions in mammalian cells. J. Mol. Appl. Gen. 2:101–109; 1983.Google Scholar
- 9.Lèvesque, J.-P.; Sansilvestri, P.; Hatzfeld, A., et al. DNA transfection in COS cells: A low-cost serum-free method compared to lipofection. Bio-Techniques 11:313–317; 1991.Google Scholar
- 14.Munro, S.; Maniatis, T. Expression cloning of the murine interferon Γ receptor cDNA. Proc. Natl. Acad. Sci. USA 86:9248–9252; 1991.Google Scholar
- 16.Sambrook, J.; Fritsch, E. F.; Maniatis, T. Molecular cloning: a laboratory manual, 2nd ed. New York: Cold Spring harbor Laboratory Press; 1989.Google Scholar