Summary
In this paper, we describe a simple technique to introduce DNA into cells through cracks and/or pores in cell membranes caused by intracellular ice crystal formation induced by liquid nitrogen. We mixed mouse BALB 3T3 cells and pSV2-neo DNA and froze the cell suspension under various conditions to determine those optimum for the introduction of DNA into mammalian cells. We found that brief treatment with liquid nitrogen, which showed only moderate cell killing, resulted in the induction of G-418 resistant colonies. These results suggest that this new technique is useful for transfection of genes into mammalian cells.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Barbacid, M.ras oncogenes in human and carcinogen-induced animal tumors. In: Klein, G., ed. Cellular oncogene activation. New York: Marcel Dekker; 1988:121–147.
Crowley, J. M. Electrical breakdown of bimolecular lipid membranes as an electromechanical instability. Biophysical J. 13:717–724; 1983.
Graessmann, M.; Graessman, A. “Early” simian-virus-40-specific RNA contains information for tumor antigen formation and chromatin replication. Proc. Natl. Acad. Sci. USA 73:366–370; 1976.
Graham, F. L.; Van der Eb, A. J. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology 52:456–467; 1973.
Iino, T.; Furusawa, M.; Furusawa, I., et al. Transformation of L cells with virus thymidine kinase genes introduced by red cell-mediated microinjection. Exp. Cell Res. 148:475–480; 1983.
Kakunaga, T.; Crow, J. D. Cell variants showing differential susceptibility to ultraviolet light-induced transformation. Science 226:552–553; 1980.
Kurata, S.; Tsukakoshi, M.; Kasuya, T., et al. The laser method for efficient introduction of foreign DNA into culture cells. Exp. Cell Res. 162:372–378; 1986.
Lavitrano, M.; Camaioni, A.; Fazio, V. M., et al. Sperm cells as vectors for introducing foreign DNA into eggs: genetic transformation of mice. Cell 57:717–723; 1989.
Meryman, H. T. Mechanisms of freezing in living cells and tissues. Science 124:515–521; 1956.
Meryman, H. T. Preservation of living cells. Fed. Proc. 22:81–89; 1963.
Mizusawa, H.; Tanaka, S.; Kobayashi, M., et al. Alkali-labile colicinogenic factor El DNA molecules formed in the presence of N-methyl-N′-nitro-N-nitrosoguanidine. Biochem. Biophys. Res. Commun. 74:570–576; 1977.
Morse, H. C. III; Rapp, U. R. Tumorigenic activation of artifically activated oncogenes. In: Klein, G., ed. Cellular oncogene activation. New York: Marcel Dekker; 1988:335–364.
Sasaki, K.; Mizusawa, H.; Ishidate, M. Isolation and characterization ofras-transfected BALB/3T3 clone showing morphological transformation by 12-O-tetradecanoyl-phorbol-13-acetate. Jap. J. Cancer Res. (Gann) 79:921–930; 1988.
Schaefer-Ridder, M.; Wang, Y.; Hofschneider, P. H. Liposomes as gene carriers: efficient transformation of mouse L cells by thymidine kinase gene. Science 215:166–168; 1982.
Schaffner, W. Direct transfer of cloned genes from bacteria to mammalian cells. Proc. Natl. Acad. Sci. USA 77:2163–2167; 1980.
Southern, P J.; Berg, P. Transformation of mammalian cells antibiotic resistance with a bacterial gene under control of the SV40 early region promoter. J. Mol. Appl. Genet. 1:327–341; 1982.
Wong, T.-K.; Neumann, E. Electric field mediated gene transfer. Biochem. Biophys. Res. Commun. 107:584–587; 1982.
Author information
Authors and Affiliations
Additional information
This work was supported by a Grant-in Aid from the Ministry of Health and Welfare for the Comprehensive 10-Year Strategy for Cancer Control, Japan.
Rights and permissions
About this article
Cite this article
Sasaki, K., Mizusawa, H., Ishidate, M. et al. Gene transfer into mammalian cells by rapid freezing. In Vitro Cell Dev Biol - Animal 27, 86–88 (1991). https://doi.org/10.1007/BF02630898
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02630898