Single-copy transgenic mice with chosen-site integration

Sarah K. Bronson, Elizabeth G. Plaehn, Kimberly D. Kluckman, John R. Hagaman, Nobuyo Maeda, Oliver Smithies

Research output: Contribution to journalArticlepeer-review

249 Scopus citations

Abstract

We describe a general way of introducing transgenes into the mouse germ line for comparing different sequences without the complications of variation in copy number and insertion site. The method uses homologous recombination in embryonic stem (ES) cells to generate mice having a single copy of a transgene integrated into a chosen location in the genome. To test the method, a single copy murine bcl-2 cDNA driven by either a chicken β-actin promoter or a human β-actin promoter has been inserted immediately 5' to the X-linked hypoxanthine phosphoribosyltransferase locus by a directly selectable homologous recombination event. The level of expression of the targeted bcl-2 transgene in ES cells is identical in independently isolated homologous recombinants having the same promoter yet varies between the different promoters. In contrast, the expression of bcl-2 transgenes having the same (chicken β-actin) promoter varies drastically when they are independently integrated at random insertion sites. Both promoters direct broad expression of the single-copy transgene in mice derived from the respective targeted ES cells. In vitro and in vivo, the human β-actin promoter consistently directed a higher level of transgene expression than the chicken β-actin promoter.

Original languageEnglish (US)
Pages (from-to)9067-9072
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume93
Issue number17
DOIs
StatePublished - Aug 20 1996

All Science Journal Classification (ASJC) codes

  • General

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