Application of novel vectors for GFP-tagging of proteins to study microtubule-associated proteins

Gene. 1996;173(1 Spec No):107-11. doi: 10.1016/0378-1119(95)00899-3.

Abstract

We describe the construction of pBact-NGFP and pBact-CGFP, two expression vectors that incorporate green fluorescent protein (GFP) as a fluorescent tag at the N- or C terminus of the produced protein. When transfected into recipient cells, GFP-tagged proteins can be visualised in the living cells using standard fluorescence microscopy techniques. Using these expression vectors, we have produced GFP-tagged versions of the neuronal microtubule-associated proteins (MAP), MAP2c and Tau34, in a number of different cell types. Both GFP-MAP2c and GFP-Tau34 were fluorescent and retained their ability to bind to microtubules. The pBact-NGFP and pBact-CGFP expression vectors represent a fast and convenient way to produce fluorescently tagged polypeptides of selected sequences encoding whole proteins or fragments for the analysis of function and dynamic events in living cells.

MeSH terms

  • Animals
  • Base Sequence
  • Cloning, Molecular
  • Cytoskeleton / metabolism
  • DNA, Complementary
  • Genetic Vectors
  • Green Fluorescent Proteins
  • HeLa Cells
  • Humans
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism*
  • Microscopy, Fluorescence
  • Microtubule-Associated Proteins / genetics
  • Microtubule-Associated Proteins / metabolism*
  • Molecular Sequence Data
  • Scyphozoa
  • Transfection

Substances

  • DNA, Complementary
  • Luminescent Proteins
  • Microtubule-Associated Proteins
  • Green Fluorescent Proteins