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Related Experiment Videos

An efficient expression, purification and immunodetection system for recombinant gene products

G Baier1, G Baier-Bitterlich, C Couture

  • 1La Jolla Institute for Allergy and Immunology, CA.

Biotechniques
|July 1, 1994
PubMed
Summary

Researchers developed a modified expression vector, pTag/CMV-neo, for easy purification and detection of recombinant fusion proteins using a novel peptide tag. This tool aids in characterizing proteins expressed from the human cytomegalovirus (CMV) or T7 promoters.

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Area of Science:

  • Molecular Biology
  • Recombinant Protein Expression

Context:

  • Mammalian expression vectors are crucial for producing recombinant proteins.
  • Current methods may lack efficiency in purification and specific detection.
  • The pRc/CMV vector offers gene expression control via CMV or T7 promoters.

Purpose:

  • To modify the pRc/CMV mammalian expression vector for enhanced recombinant fusion protein expression.
  • To enable simple, single-step affinity purification and specific immunodetection of fusion proteins.
  • To introduce a novel peptide tag for improved protein characterization.

Summary:

  • A modified mammalian expression vector, pTag/CMV-neo, was engineered from pRc/CMV.
  • It incorporates a Kozak consensus ribosome-binding site and a 30-amino acid fusion tag.

Related Experiment Videos

  • The tag includes a (His)6 metal-binding site for Ni(2+)-chelating resin purification and a p18HIV peptide for immunodetection with monoclonal antibody H902.

    • Impact:

    • Facilitates straightforward purification and specific detection of recombinant fusion proteins.
    • The p18HIV epitope and H902 antibody offer a specific tool for protein identification and characterization.
    • This modified vector system enhances the utility of expression systems driven by CMV or T7 promoters.