Abstract
Retroviral cDNA expression libraries allow the efficient introduction of complex cDNA libraries into virtually any mitotic cell type for screening based on gene function. The cDNA copy number per cell can be easily controlled by adjusting the multiplicity of infection, thus cell populations may be generated in which >90% of infected cells contain one to three cDNAs. We describe the isolation of two known oncogenes and one cell-surface receptor from a human Burkitt’s lymphoma (Daudi) cDNA library inserted into the hightiter retroviral vector pFB.
Similar content being viewed by others
References
Kitamura, T., Onishi, M., Kinoshita, S., Shibuya, A., Miyajima, A., and Nolan, G. P. (1995) Efficient screening of retroviral cDNA expression libraries. Proc. Natl. Acad. Sci. 92, 9146–9150.
Kitamura, T. (1998) New experimental approaches in retrovirus-mediated expression screening. Int. J. Hematol. 67, 351–359.
Onishi, M., Kinoshita, S., Morikawa, Y., et al. (1996) Applications of retrovirus-mediated expression cloning. Exp. Hematol. 24, 324–329.
Rayner, J. R. and Gonda, T. J. (1994) A simple and efficient procedure for generating stable expression libraries by cDNA cloning in a retroviral vector. Mol. Cell Biol. 14, 880–887.
Miller, A. D. (1997) Retroviruses (Coffin, J. M., Hughes, S. H., and Varmus, H. E., eds.), Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, pp. 437–473.
Riviere, I. and Sadelain, M. (1997) Methods in Molecular Medicine: Gene Therapy Protocols (Robbins, P.D., ed.), Humana Press, Totowa, NJ, p. 59–78.
Vaillancourt, P., Rogers, B. J., Chen, K., and Felts, K. A. (2002) Transient Production of high-titer retroviral vectors for high-throughput applications, in Gene Cloning and Expression Technologies (Lu, Q. and Weiner, M. P., eds.) BioTechniques Books, Eaton Publishing, Natick, MA, pp. 1–13.
Naviaux, R. K., Costanzi, E., Haas, M. and Verma, I. M. (1996) The pCL vector system: rapid production of helper-free, high-titer, recombinant retroviruses. J. Virol. 70, 5701–5705.
Pear, W. S., scott, M. L., and Nolan, G. P. (1997) Generation of high-titer, helper-free retroviruses by transient transfection, in Methods in Molecular Medicine: Gene Therapy Protocols (Robbins, P. D., ed.), Humana Press, Totowa, NJ, pp. 41–58.
Pages, J. C., Loux, N., Farge, D., Briand, P., and Weber, A. (1995) Activation of Moloney murine leukemia virus LTR enhances the titer of recombinant retrovirus in psi CRIP packaging cells. Gene Ther. 8, 547–551.
Riviere, I., Brose, K., and Mulligan, R. C. (1995) Effects of retroviral vector design on expression of human adenosine deaminase in murine bone marrow transplant recipients engrafted with genetically modified cells. Proc. Natl. Acad. Sci. USA 92, 6733–6737.
Hesketh, R. (1997) The Oncogene and Tumour Suppressor Gene FactsBook, Academic Press, San Diego.
Lorenzi, M. V., Long, J. E., Miki, T., and Aaronson, S. A. (1995) Expression cloning, developmental expression and chromosomal localization of fibroblast growth factor-8. Oncogene 10, 2051–2055.
Lorenzi, M. V., Horii, Y., Yamanaka, R., Sakaguchi, K., and Miki, T. (1996) FRAG1, a gene that potently activates fibroblast growth factor receptor by C-terminal fusion through chromosomal rearrangement. Proc. Natl. Acad. Sci. USA 93, 8956–8961.
Whitehead, I., Kirk, H., and Kay, R. (1995) Expression cloning of oncogenes by retroviral transfer of cDNA libraries. Mol. Cell Biol. 15, 704–710.
Ren, S., Smith, M. J., Louro, I. D., et al. (2000) The p44S10 locus, encoding a subunit of the proteasome regulatory particle, is amplified during progression of cutaneous malignant melanoma. Oncogene 19, 1419–1427.
Tatsuka, M., Mitsui, H., Wada, M., Nagata, A., Nojima, H., and Okayama, H. (1992) Elongation factor-1 alpha gene determines susceptibility to transformation. Nature 359, 333–336.
Liu, Y., Wei, S. H., Ho, A. S., de Waal Malefyt, R., and Moore, K. W. (1994) Expression cloning and characterization of a human IL-10 receptor. J. Immunol. 152, 1821–1829.
Ikizawa, K., Kajiwara, K., Koshio, T., Matsuura, N., and Yanagihara, Y. (1995) Inhibition of IL-4 receptor up-regulation on B cells by antisense oligodeoxynucleotide suppresses IL-4-induced human IgE production. Clin. Exp. Immunol. 3, 383–389.
Brooks, D. G., Qiu, W. Q., Luster, A. D., and Ravetch, J. V. (1989) Structure and expression of human IgG FcRII(CD32). Functional heterogeneity is encoded by the alternatively spliced products of multiple genes. J. Exp. Med. 170, 1369–1385.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Felts, K.A., Chen, K., Zaharee, K. et al. Functional cloning using pFB retroviral cDNA expression libraries. Mol Biotechnol 22, 25–32 (2002). https://doi.org/10.1385/MB:22:1:025
Issue Date:
DOI: https://doi.org/10.1385/MB:22:1:025