It was found in our previous study (Oda et al., 1990. J. Biol. Chem. 265: 7513-7519) that in the rat two mRNAs encoding mitochondrial and peroxisomal serine:pyruvate aminotransferase (SPT/AGT) are formed from a single SPT/AGT gene through alternative transcription initiation in exon 1. In an attempt to analyze the mechanisms underlying this unique phenomenon, we have isolated genomic clones harboring the entire rat SPT/AGT gene. In the present study, the location of the rat SPT/AGT gene was determined to be in the q34-q36 region of chromosome 9 by fluorescence in situ hybridization. Southern blot analysis of rat genomic DNA revealed an allelic BamHI restriction fragment length polymorphism among three different inbred rat strains. These results indicated that a single copy SPT/AGT gene is located on chromosome 9q34-q36 in the rat genome. This locus has been assigned the gene symbol Spat.