Retroviral vectors for establishing tetracycline-regulated gene expression in an otherwise recalcitrant cell line

Paraic A. Kenny, Tariq Enver, Alan Ashworth

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Background: Tetracycline-regulated systems have been used to control the expression of heterologous genes in such diverse organisms as yeast, plants, flies and mice. Adaptation of this prokaryotic regulatory system avoids many of the problems inherent in other inducible systems. There have, however, been many reports of difficulties in establishing functioning stable cell lines due to the cytotoxic effects of expressing high levels of the tetracycline transactivator, tTA, from a strong viral promoter. Results: Here we report the successful incorporation of tetracycline-mediated gene expression in a mouse mammary epithelial cell line, HC11, in which conventional approaches failed. We generated retroviruses in which tTA expression was controlled by one of three promoters: a synthetic tetracycline responsive promoter (TRE), the elongation factor 1-alpha promoter (EF1α) or the phosphoglycerate kinase-1 promoter (PGK), and compared the resulting cell lines to one generated using a cytomegalovirus immediate early gene promoter (CMV). In contrast to cells produced using the CMV and PGK promoters, those produced using the EF1α and TRE promoters expressed high levels of β-galactosidase in a tetracycline-dependent manner. Conclusions: These novel retroviral vectors performed better than the commercially available system and may have a more general utility in similarly recalcitrant cell lines.

Original languageEnglish (US)
Article number13
JournalBMC Molecular Biology
Volume3
DOIs
StatePublished - Sep 3 2002
Externally publishedYes

Fingerprint

Tetracycline
Gene expression
Cells
Gene Expression
Cell Line
Phosphoglycerate Kinase
Peptide Elongation Factor 1
Genes
Galactosidases
Immediate-Early Genes
Trans-Activators
Retroviridae
Cytomegalovirus
Diptera
Yeast
Breast
Yeasts
Epithelial Cells

Keywords

  • Doxycycline
  • Inducible expression
  • Retroviral vectors
  • Tetracyline
  • Transgenics

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry

Cite this

Retroviral vectors for establishing tetracycline-regulated gene expression in an otherwise recalcitrant cell line. / Kenny, Paraic A.; Enver, Tariq; Ashworth, Alan.

In: BMC Molecular Biology, Vol. 3, 13, 03.09.2002.

Research output: Contribution to journalArticle

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