DsRed-Express, a popular reporter protein, cannot be expressed in Escherichia coli using a consensus ribosome binding site (RBS) potentially due to base-pairing in the RBS that inhibits translation initiation. Saturation mutagenesis was used to probe for a gene sequence that minimized base-pairing in the RBS while maintaining the same spectral properties and maturation characteristics as DsRed-Express. The new DsRed, designated here as RFPEC for E. coli optimized red fluorescent protein, fluoresces 2.5 times greater than DsRed-Express when expressed from the same vector. (c) 2005 Wiley Periodicals, Inc.