A breakthrough in the availability of spectrally distinct autofluorescent proteins came with the cloning of six anthozoan fluorescent proteins all having 20–30% identity to wtGFP [12]. Of these, DsRed1 (referred to as drFP585 by Matz et al., [12]), a 28 kDa protein isolated from the IndoPacific sea anemone Discosoma sp, posseses the longest excitation (558 nm) and emission (583 nm) wavelength maxima known for a wild-type autofluorescent protein. The sequence of DsRed1 has recently been modified and optimized for high levels of expression in mammalian cells by the introduction of 144 silent base pair changes corresponding to human codon usage preferences. We therefore also tested this novel red fluorescent protein (RFP) in ES cells and mice.