Different national and international agencies have selected specific STR sets for forensic database use. To enhance database comparison across national and international borders, a 27-locus multiplex system was developed comprising all 15 STR loci of the European standard set, the current 13 STR loci of the CODIS core, the proposed 22 STR loci of the expanded CODIS core, 4 additional commonly used STR loci, and the amelogenin locus. Development required iterative primer design to resolve primer-related artifacts, amplicon sizing, and locus-to-locus balance issues. The 19.5-min assay incorporated newly developed six-dye chemistry analyzed using a novel microfluidic electrophoresis instrument capable of simultaneous detection and discrimination of 8 or more fluorescent dyes. The 27-locus multiplex offers the potential for a new international STR standard permitting laboratories in any jurisdiction to use a single reaction to determine profiles for loci they typically generate plus an expanded common STR profiling set of global interest.