Excerpts

In HEK293 cells expressing monomeric eqFP611 variants, the red fluorescence was not evenly distributed but rather appeared as dot-like structures in the cytoplasm... A similar effect was noticed for eqFP611 dimers that had the A/C interface disrupted. This result suggested that exposure of the A/C interface may have made a subcellular targeting signal accessible. Indeed, sequence analysis revealed that amino acids 229GRL231 at the C terminus, and also the preceding triplet 226SKL228 may serve as a type-1 peroxisomal targeting signal... We succeeded in removing peroxisomal targeting by replacing the C-terminal sequence 222CDLPSKLGRL231 of eqFP611 by 222AGLGGG227 [in mRuby]

Analysed by flow cytometry, the fluorescence signal of living HEK293 cells expressing the codon optimized variant [of mRuby] was 5–8 fold increased in comparison to the predecessor with unaltered codon usage but identical amino acid sequence.