Summary
The use of polybrene for the efficient transfection of cultured mosquito cells is described. Cells are transfected with purified plasmid DNA coding for a bacterial chloramphenicol acetyltransferase (CAT) gene regulated by sequences from aDrosophila heat shock protein promoter. The DNA is added to the cells in serumfree medium containing polybrene, which facilitates adsorption of DNA to the cell surface. Expression of the transfected gene is induced by subjecting recipient cells to heat shock conditions. CAT activity in lysates from transfected cells is assayed using a simple thin layer chromatographic procedure. Adaptation of the transfection protocol to other insect cell types is discussed.
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References
Clewell, D. B.; Helinski, D. R. Properties of a supercoiled deoxyribonucleic acid-protein relaxation complex and strand specificity of the relaxation event. Biochemistry 9:4429–4440; 1970.
Corces, V.; Pellicer, A.; Axel, R., et al. Integration, transcription and control of aDrosophila heat shock gene in mouse cells. Proc. Natl. Acad. Sci. USA 78:7038–7042; 1981.
Di Nocera, P. P; Dawid, I. B. Transient expression of genes introduced into cultured cells ofDrosophila. Proc. Natl. Acad. Sci. USA 80:7095–7098; 1983.
Dulbecco, R.; Vogt. M. Plaque formation and isolation of pure lines with poliomyelitis viruses. J. Exp. Med. 99:167–182; 1954.
Durbin, J. E.; Fallon, A. M. Transient expression of the chloramphenicol acetyltransferase gene in cultured mosquito cells. Gene 36:173–178; 1985.
Fallon, A. M. Factors affecting polybrene-mediated transfection of culturedAedes albopictus (mosquito) cells. Exp. Cell Res. 166:535–542; 1986.
Fallon, A. M.; Stollar, V. The biochemistry and genetics of mosquito cells in culture. In: Maramorosch, K., ed. Advances in cell culture, vol. 5. New York: Academic Press; 1987:97–137.
Lowe, D. G.; Fulford, W. D.; Moran, L. A. Mouse andDrosophila genes encoding the major heat shock protein (hsp 70) are highly conserved. Mol. Cell. Biol. 3:1540–1543; 1983.
Maniatis, T.; Fritsch, E. F.; Sambrook, J. Molecular cloning: a laboratory manual. New York: Cold Spring Harbor Laboratory; 1982.
Swerdel, M. R.; Fallon, A. M. Phosphoribosylation of xanthine by extracts from insect cells. Insect Biochem. 17:1181–1186; 1987.
Voellmy, R.; Rungger, D. Transcription of aDrosophila heat shock gene is heat-induced inXenopus occytes. Proc. Natl. Acad. Sci. USA 79:1776–1780; 1982.
Walker, V. K. Gene transfer in insects. In: Maramorosch, K.; Sato, G., eds. Advances in cell culture, vol. 7. New York: Academic Press; in press, 1989.
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Fallon, A.M. Optimization of gene transfer in cultured insect cells. Journal of Tissue Culture Methods 12, 1–6 (1989). https://doi.org/10.1007/BF01577999
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DOI: https://doi.org/10.1007/BF01577999