Genetic mapping of the structural gene for a phenylalanyl-sRNA synthetase in Escherichia coli was accomplished by the use of mutants with temperature-sensitive or p-fluorophenylalanine-resistant enzymes. The structural gene is located at minute 33 of the Taylor-Thoman map of the E. coli chromosome, closely linked to a structural gene (aroD) for one of the enzymes involved in the biosynthesis of the aromatic amino acids, and distant from the known locations of other aminoacyl-sRNA synthetase genes.