Variant Forms of Green and BIue Fluorescent Proteins Adapted for the Use in Mammalian Cells

• Yamasaki Masatosi

  Department of Life Science, Himeji Institute of Technology

• Hashiguchi Noriyo

  Department of Life Science, Himeji Institute of Technology

• Tsukamoto Toshiro

  Department of Life Science, Himeji Institute of Technology

• Osumi Takashi

  Department of Life Science, Himeji Institute of Technology

Abstract

Green fluorescent protein (GFP) and its variants are highly convenient markers to trace the movement of proteins in living cells. Their fluorescence, however, is not strong enough at physiological temperatures for mammalian cells, and the blue variant, blue fluorescent protein (BFP), is photo-bleached quickly. To overcome these limitations, we introduced random mutations to the BFP sequence by mutagenic PCR, and selected clones that fluoresce brighter, after prolonged exposure to UV light. One of such clones had an amino acid substitution adjacent to the chromophore. By combining this mutation with mutations known to improve the fluorescence intensity, we obtained GFP and BFP variants having remarkably stronger fluorescence when expressed in mammalian cells as well as in Eschelichia coli. These mutants have increased stability and are present in the active species in higher proportions at 37°C than are the original proteins. Thus, they would be highly convenient tools for mammalian cell biology.

Journal

• bioimages

  bioimages 6 (1), 1-7, 1998

  Bioimaging Society

Keywords

• Green fluorescent protein
• blue fluorescent protein
• GFP
• BFP
• genetic engineering

Details

• CRID: 1390001277364306176
• NII Article ID: 10002038405
• NII Book ID: AA11084187
• DOI: 10.11169/bioimages.6.1
• ISSN: 09192719
• Text Lang: en
• Data Source:
  • JaLC
  • CiNii Articles
• Abstract License Flag: Disallowed