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DNA-mediated gene transfer into epidermal cells using electroporation.

M Reiss, M M Jastreboff, J R Bertino

    Biochemical and Biophysical Research Communications
    |May 29, 1986
    PubMed
    Summary

    Researchers developed a reliable method for introducing foreign DNA into epidermal cells using electroporation. This technique successfully demonstrated transient gene expression and stable DNA integration in mouse cells.

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    Area of Science:

    • Molecular Biology
    • Cell Biology
    • Genetics

    Background:

    • Efficient delivery of foreign DNA into epidermal cells is crucial for genetic studies and therapeutic applications.
    • Electroporation offers a potential non-viral method for gene transfer into various cell types.

    Purpose of the Study:

    • To establish and validate a reliable electroporation-based method for introducing foreign DNA into murine epidermal cells (MK-1).
    • To assess the efficiency of transient gene expression and stable DNA integration following electroporation.

    Main Methods:

    • Murine BALB/c MK-1 epidermal cells were subjected to electroporation with plasmid DNA constructs (pSV2-CAT and pSV2-neo).
    • Transient expression of chloramphenicol-acetyltransferase (CAT) was quantified.
    • Selection for G418 resistant clones was performed after transfection with pSV2-neo.

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  • Integration patterns of the SV2-neo construct were analyzed in resistant clones.

    • Main Results:

    • Electroporation resulted in transient expression of chloramphenicol-acetyltransferase, with activity ranging from 0.03 to 0.05 nmoles acetylchloramphenicol per mg protein per min.
    • Transfection with pSV2-neo yielded approximately eight G418 resistant clones per 10(-6) cells per microgram of plasmid DNA.
    • Analysis revealed distinct integration patterns of the SV2-neo construct in three independently selected resistant clones.

    Conclusions:

    • Electroporation is an effective method for introducing foreign DNA into murine epidermal cells.
    • The established method allows for both transient gene expression and stable integration of foreign DNA.
    • This technique provides a valuable tool for genetic manipulation of epidermal cells.