Expression of the lacZ gene targeted to the HPRT locus in embryonic stem cells and their derivatives

Jessica R. Shaw-white, Nicholas Denko, Lisa Albers, Thomas C. Doetschman, James R. Stringer

Research output: Contribution to journalArticlepeer-review

17 Scopus citations

Abstract

Transgenes in mice often exhibit different expression patterns in different transgenic lines. While the basis for this phenomenon is not understood, it is widely believed that the site at which the transgene becomes integrated into the mouse genome is a major factor in determining the pattern of expression. Most transgenic mice have been produced by microinjection of DNA into the male pronucleus, which results in integration of tandem arrays of the transgene at random chromosomal sites. In the experiments described in this report, electroporation of embryonic stem (ES) cells was used to place single copies of a lacZ transgene into either random sites or into the HPRT (hypoxanthine phosphoribosyl transferase) locus of the mouse genome. Expression of lacZ was assayed by histochemical staining for Escherichia coli β-galactosidase activity in ES cells and in differentiated derivatives obtained by teratocarcinoma formation. Several of the randomly integrated cell lines expressed lacZ at high levels in a variety of cell types present in the tumours, but most notably in epithelial cells. Targeted cell lines with lacZ in opposite orientation to the direction of HPRT gene transcription also expressed well in epithelial cells, but the targeted cell lines did not express in a wider variety of cell types than some of the nontargeted cell lines. Targeted cell lines transcribing lacZ in the same orientation as HPRT transcription did not express high levels of lacZ in any differentiated cell type. Analysis of transcripts suggested that this orientation effect may have been the result of transcriptional interference perpetrated by the HPRT gene promoter.

Original languageEnglish (US)
Pages (from-to)1-13
Number of pages13
JournalTransgenic Research
Volume2
Issue number1
DOIs
StatePublished - Jan 1993

Keywords

  • ES cells
  • HPRT
  • gene expression
  • gene targeting
  • lacZ

ASJC Scopus subject areas

  • Biotechnology
  • Animal Science and Zoology
  • Agronomy and Crop Science
  • Genetics

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