Abstract We have employed a new approach to generate novel fluorescent proteins (FPs) from red absorbing chromoproteins. An identical single amino acid substitution converted novel chromoproteins from the species Anthozoa ( Heteractis crispa, Condylactis gigantea, and Goniopora tenuidens) into far-red FPs (emission λ max=615–640 nm). Moreover, coupled site-directed and random mutagenesis of the chromoprotein from H. crispa resulted in a unique far-red FP (HcRed) that exhibited bright emission at 645 nm. A clear red shift in fluorescence of HcRed, compared to drFP583 (by more than 60 nm), makes it an ideal additional color for multi-color labeling. Importantly, HcRed is excitable by 600 nm dye laser, thus promoting new detection channels for multi-color flow cytometry applications. In addition, we generated a dimeric mutant with similar maturation and spectral properties to tetrameric HcRed.