The uricase gene was isolated from rat genomic DNA libraries. The gene spans 40 kb and consists of eight exons. All the exon-intron junctional sequences conform to the canonical GT/AG rule. The restriction map of the isolated clones and Southern blot analysis revealed that the enzyme is encoded by a single-copy gene. Analysis of the transcription initiation site of rat uricase mRNA indicated the differential use of consecutive nucleotides; the principal repeat is located 55 nucleotides upstream from the first methionine codon. Nucleotide sequence analysis of the 5'-flanking region showed the presence of a TATA (ATAAAA) sequence at nucleotides 30 to 25 and of a CAAT (GGTCAAT) sequence at nucleotides 63 to 57 upstream of the principal transcription initiation site. The 5'-flanking region contains another possible regulatory sequence (TGTCGACA) homologous to the cAMP-regulatory element. The palindromic sequence is located 158 to 151 nucleotides upstream from the transcription initiation site, surrounded by a direct repeat (TCAGCAA).