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A heat-shock-inducible eukaryotic expression vector.

C W Schweinfest1, C L Jorcyk, S Fujiwara

  • 1Laboratory of Molecular Oncology, National Cancer Institute, Frederick, MD 21701-1013.

Gene
|November 15, 1988
PubMed
Summary
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Researchers developed pHS/Cla, an inducible expression vector using the heat shock protein 70 (hsp70) promoter from Drosophila melanogaster. This vector allows heat-inducible expression of genes, such as the human immunodeficiency virus (HIV) transactivator, in mammalian cells with tight regulation.

Area of Science:

  • Molecular Biology
  • Gene Expression
  • Virology

Background:

  • Development of inducible expression systems is crucial for controlling gene function in mammalian cells.
  • The heat shock protein 70 (hsp70) promoter from Drosophila melanogaster is a well-characterized inducible promoter.
  • Human immunodeficiency virus (HIV) transactivator protein is a key factor in viral gene expression.

Purpose of the Study:

  • To create a novel inducible expression vector, pHS/Cla, utilizing the Drosophila melanogaster hsp70 promoter.
  • To demonstrate the functional expression of the HIV-1 transactivator protein in mammalian cells using this vector.
  • To characterize the inducibility and regulation of the hsp70 promoter in the pHS/Cla construct.

Main Methods:

  • Construction of the pHS/Cla plasmid, based on pAT153, incorporating the Drosophila melanogaster hsp70 promoter.

Related Experiment Videos

  • Transient transfection of human H9 T-cells with the pHS/Cla construct.
  • Analysis of gene expression in response to heat shock conditions.

    • Main Results:

    • The pHS/Cla construct successfully facilitated heat-inducible expression of the HIV-1 transactivator protein.
    • The hsp70 promoter demonstrated tight regulation, with no detectable expression at 37 degrees C.
    • Expression was significantly induced by heat shock and sustained for at least 4 hours.

    Conclusions:

    • The pHS/Cla construct serves as an effective inducible expression vector in mammalian cells.
    • The Drosophila melanogaster hsp70 promoter provides robust and tightly regulated gene expression upon heat induction.
    • The unique ClaI site in pHS/Cla allows for versatile cloning and expression of various genes.