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Chimeric vector construction for higher-plant transformation.

E Balázs, S Bouzoubaa, H Guilley

    Gene
    |January 1, 1985
    PubMed
    Summary
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    Researchers developed a new plant transformation vector, pKR612B1, enabling kanamycin resistance in transformed plants. This vector facilitates gene insertion and regeneration of whole plants carrying the foreign neomycin phosphotransferase gene.

    Area of Science:

    • Plant molecular biology
    • Genetic engineering
    • Biotechnology

    Background:

    • Development of efficient gene transfer systems is crucial for plant genetic modification.
    • The neomycin phosphotransferase (APH) gene confers resistance to kanamycin, a widely used selection agent.
    • Cauliflower mosaic virus (CaMV) gene VI promoter offers a strong expression system in plants.

    Purpose of the Study:

    • To construct and characterize a novel chimeric plant transformation vector, pKR612B1.
    • To evaluate the efficacy of pKR612B1 for transforming higher plant protoplasts.
    • To regenerate kanamycin-resistant plants expressing the introduced foreign gene.

    Main Methods:

    • Construction of the chimeric vector pKR612B1 containing the Tn5 APH gene under CaMV gene VI promoter control.

    Related Experiment Videos

  • Transformation of turnip and tobacco protoplasts using the polyethylene glycol (PEG) method.
  • Regeneration of whole plants from transformed protoplasts and molecular analysis (Southern blot, enzyme activity assays).

    • Main Results:

    • Successful transformation of turnip and tobacco protoplasts, conferring kanamycin resistance.
    • Regeneration of whole, kanamycin-resistant tobacco plants from transformed protoplasts.
    • Confirmation of integrated foreign gene presence and functional neomycin phosphotransferase (APH) activity in calli and regenerated plants.

    Conclusions:

    • The developed vector pKR612B1 is effective for introducing foreign genes into higher plants.
    • The vector facilitates the regeneration of genetically modified plants with selectable marker expression.
    • This system provides a valuable tool for plant genetic engineering and functional genomics studies.