Insertion of a foreign gene into the beta-casein locus by Cre-mediated site-specific recombination

Andreas Kolb, R Ansell, J McWhir, S G Siddell

    Research output: Contribution to journalArticle

    34 Citations (Scopus)

    Abstract

    The expression of foreign genes in transgenic animals is generally unpredictable as transgenes are integrated at random after pro-nuclear injection into fertilized oocytes. In many cases, transgene expression is inhibited by neighbouring chromatin structures or by the repeated nature of the multiple transgene copies present at the integration site. A strategy involving homologous and site-specific recombination has been devised by which single copies of a foreign gene can be inserted specifically into the locus of a highly expressed gene. As a first step, a loxP recombination target site is introduced by homologous recombination into a predetermined gene locus such that the loxP sequence is placed next to the promoter region and replaces the translational initiation signal. In a subsequent site-specific recombination reaction, a gene of interest can be integrated into the pre-existing loxP site. This biphasic recombination strategy was used to integrate a luciferase reporter gene into the locus of the murine beta-casein gene in embryonic stem cells. (C) 1999 Elsevier Science B.V. All rights reserved.

    Original languageEnglish
    Pages (from-to)21-31
    Number of pages11
    JournalGene
    Volume227
    Issue number1
    DOIs
    Publication statusPublished - 4 Feb 1999

    Keywords

    • embryonic stem cells
    • gene expression
    • genome engineering
    • homologous recombination
    • embryonic stem-cells
    • alpha-lactaalbumin gene
    • transgenic mice
    • control region
    • globin genes
    • mammalian-cells
    • tyrosinase gene
    • expression
    • replacement
    • protein

    Cite this

    Insertion of a foreign gene into the beta-casein locus by Cre-mediated site-specific recombination. / Kolb, Andreas; Ansell, R ; McWhir, J ; Siddell, S G .

    In: Gene, Vol. 227, No. 1, 04.02.1999, p. 21-31.

    Research output: Contribution to journalArticle

    Kolb, Andreas ; Ansell, R ; McWhir, J ; Siddell, S G . / Insertion of a foreign gene into the beta-casein locus by Cre-mediated site-specific recombination. In: Gene. 1999 ; Vol. 227, No. 1. pp. 21-31.
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    AB - The expression of foreign genes in transgenic animals is generally unpredictable as transgenes are integrated at random after pro-nuclear injection into fertilized oocytes. In many cases, transgene expression is inhibited by neighbouring chromatin structures or by the repeated nature of the multiple transgene copies present at the integration site. A strategy involving homologous and site-specific recombination has been devised by which single copies of a foreign gene can be inserted specifically into the locus of a highly expressed gene. As a first step, a loxP recombination target site is introduced by homologous recombination into a predetermined gene locus such that the loxP sequence is placed next to the promoter region and replaces the translational initiation signal. In a subsequent site-specific recombination reaction, a gene of interest can be integrated into the pre-existing loxP site. This biphasic recombination strategy was used to integrate a luciferase reporter gene into the locus of the murine beta-casein gene in embryonic stem cells. (C) 1999 Elsevier Science B.V. All rights reserved.

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    KW - genome engineering

    KW - homologous recombination

    KW - embryonic stem-cells

    KW - alpha-lactaalbumin gene

    KW - transgenic mice

    KW - control region

    KW - globin genes

    KW - mammalian-cells

    KW - tyrosinase gene

    KW - expression

    KW - replacement

    KW - protein

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