Far-red fluorescent tag for protein labelling

Practical applications of green fluorescent protein (9GFP9)-like fluorescent proteins (FPs) from species of the class Anthozoa (sea anemones, corals and sea pens) are strongly restricted owing to their oligomeric nature. Here we suggest a strategy to overcome this problem by the use of two covalently linked identical red FPs as non-oligomerizing fusion tags. We have applied this approach to the dimeric far-red fluorescent protein HcRed1 and have demonstrated superiority of the tandem tag in the in vivo labelling of fine cytoskeletal structures and tiny nucleoli. In addition, a possibility of effective fluorescence resonance energy transfer (9FRET9) between enhanced yellow FP mutant (9EYFP9) and tandem HcRed1 was demonstrated in a protease assay.