Plasmids for the cloning and expression of full-length double-stranded cDNAs under control of the SV40 early or late gene promoter - PubMed

Plasmids for the cloning and expression of full-length double-stranded cDNAs under control of the SV40 early or late gene promoter

R Breathnach et al. Nucleic Acids Res. 1983.

Free PMC article

Abstract

Okayama and Berg (1) have recently described a technique for the high efficiency cloning of full-length dscDNAs. We have constructed eukaryotic expression vectors compatible both with this technique (and with classical techniques) for dscDNA cloning. The vectors are such that recombinants obtained contain dscDNAs in the correct orientation downstream from a block of sequence comprising either the SV40 early or late gene promoter linked to a pair of splice sites from a rabbit beta-globin gene. A sequence encoding an SV40 polyadenylation site follows the dscDNA. We have used our vectors to make a library from chicken oviduct polyA(+) RNA using the Okayama and Berg technique. Ovalbumin recombinants occur in the library at the expected frequency and a high proportion contain full length copies of the ovalbumin mRNA. However, a similar result was not obtained for conalbumin recombinants. When recombinants are introduced into eukaryotic cells by either calcium phosphate coprecipitation or protoplast fusion, expression of chicken ovalbumin or conalbumin may be detected by indirect immunofluorescence. Under optimal conditions (use of SV40 late promoter and cos 7 cells) ovalbumin protein could be detected when the ovalbumin recombinant was present in only 2% of the protoplasts used for fusion. This suggests that colony banks obtained using our vectors could be screened in batches of 50 by protoplast fusion followed by a search for expression of a given protein using indirect immunofluorescence.

Similar articles

• Simian virus 40 late promoter region able to initiate simian virus 40 early gene transcription in the absence of the simian virus 40 origin sequence.

  Ernoult-Lange M, May P, Moreau P, May E. Ernoult-Lange M, et al. J Virol. 1984 Apr;50(1):163-73. doi: 10.1128/JVI.50.1.163-173.1984. J Virol. 1984. PMID: 6321788 Free PMC article.

• Definition of the ovalbumin gene promoter by transfer of an ovalglobin fusion gene into cultured cells.

  Knoll BJ, Zarucki-Schulz T, Dean DC, O'Malley BW. Knoll BJ, et al. Nucleic Acids Res. 1983 Oct 11;11(19):6733-54. doi: 10.1093/nar/11.19.6733. Nucleic Acids Res. 1983. PMID: 6314256 Free PMC article.

• Construction of an adenovirus-SV40 recombinant producing SV40 T antigen from an adenovirus late promoter.

  Solnick D. Solnick D. Cell. 1981 Apr;24(1):135-43. doi: 10.1016/0092-8674(81)90509-2. Cell. 1981. PMID: 6263482

• Promoter elements of genes coding for proteins and modulation of transcription by estrogens and progesterone.

  Chambon P, Dierich A, Gaub MP, Jakowlev S, Jongstra J, Krust A, LePennec JP, Oudet P, Reudelhuber T. Chambon P, et al. Recent Prog Horm Res. 1984;40:1-42. doi: 10.1016/b978-0-12-571140-1.50005-0. Recent Prog Horm Res. 1984. PMID: 6091191 Review. No abstract available.

• SV40 virus expression vectors.

  Naim HY, Roth MG. Naim HY, et al. Methods Cell Biol. 1994;43 Pt A:113-36. doi: 10.1016/s0091-679x(08)60601-9. Methods Cell Biol. 1994. PMID: 7823859 Review.

Cited by

• Definition of an Ets1 protein domain required for nuclear localization in cells and DNA-binding activity in vitro.

  Boulukos KE, Pognonec P, Rabault B, Begue A, Ghysdael J. Boulukos KE, et al. Mol Cell Biol. 1989 Dec;9(12):5718-21. doi: 10.1128/mcb.9.12.5718-5721.1989. Mol Cell Biol. 1989. PMID: 2555704 Free PMC article.

• Epidermal growth factor or serum stimulation of rat fibroblasts induces an elevation in mRNA levels for lactate dehydrogenase and other glycolytic enzymes.

  Matrisian LM, Rautmann G, Magun BE, Breathnach R. Matrisian LM, et al. Nucleic Acids Res. 1985 Feb 11;13(3):711-26. doi: 10.1093/nar/13.3.711. Nucleic Acids Res. 1985. PMID: 3873645 Free PMC article.

• The chicken progesterone receptor: sequence, expression and functional analysis.

  Gronemeyer H, Turcotte B, Quirin-Stricker C, Bocquel MT, Meyer ME, Krozowski Z, Jeltsch JM, Lerouge T, Garnier JM, Chambon P. Gronemeyer H, et al. EMBO J. 1987 Dec 20;6(13):3985-94. doi: 10.1002/j.1460-2075.1987.tb02741.x. EMBO J. 1987. PMID: 3443098 Free PMC article.

• A primer vector system that allows temperature dependent gene amplification and expression in mammalian cells: regulation of the influenza virus NS1 gene expression.

  Portela A, Melero JA, Martínez C, Domingo E, Ortín J. Portela A, et al. Nucleic Acids Res. 1985 Nov 25;13(22):7959-77. doi: 10.1093/nar/13.22.7959. Nucleic Acids Res. 1985. PMID: 2933634 Free PMC article.

References

Publication types

MeSH terms

Substances

LinkOut - more resources

• Full Text Sources

  • Europe PubMed Central
  • PubMed Central
  • Silverchair Information Systems

• Other Literature Sources

  • The Lens - Patent Citations Database