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    The use of the mouse APRT gene in cultured human cells to study point mutations

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    Authors
    Jarrett, Robert A
    Issue Date
    1988-08
    URI

    http://hdl.handle.net/10675.2/622431
    
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    Abstract
    Site-specific mutagenesis, gene transfer, and somatic cell selection have been used to develop a method for the identification of site- and sequence-specific point mutationsinduced in a mammalian cell transgenic system. This method utilizes a mouse adenine phosphoribosyltransferase [APRT] gene containing a specific point mutation in an intronjexon splice recognition sequence, which disrupts mRNA processing and destroys a diagnostic Pst I restriction site. This construct was introduced by cotransfection,with the bacterial n&Qr gene into HTD-114, a non-spontaneously-reverting, Aprt- human fibrosarcoma-derived cell line. The construct s·tudied contains I an A to G transition which is revertible with ethyl methanesulfonate [EMS], a mutagen known to produce G to A transitions in eukaryotes and prokaryotes. Exposing two independently derived cell lines containing the construct to EMS concentrations ranging from 0 to 200 ugjml (corresponding-to 100 to 0.8 percent survival) produced a frequency of reversion to the Aprt+ phenotype of 10-7 to 10-2 .
    Affiliation
    Department of Cell and Molecular Biology
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    Theses and Dissertations

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