The isolation and characterization of the complete gene coding for human protein kinase CK2 catalytic subunit alpha is described. The gene spans 70 kb and consists of 13 exons, and the exon/intron boundaries conform to the gt/ag rule. Exons range in size from 51 to 2960 bp, introns from 527 to around 34000 bp. The translation start site is located in Exon 2, the stop codon in Exon 13. Two transcription start sites were identified by primer extension analysis: The further 5'-located site defines position 1 of the gene, the second site is located at position 50. The 5' region of the CK2 alpha gene shows features of a housekeeping promoter, such as lack of a TATA box and presence of a CpG island and GC boxes. The region was analyzed by reporter gene assay, and promoter activity was detected within the region ranging from position -256 to 144. Six potential polyadenylation signals were identified in the 3' noncoding region of the CK2 alpha gene. As indicated by comparison with expressed sequence tags from the EMBL databank and by Northern-blot analysis, the most 3' located, active polyadenylation signal seems to be the fourth signal, defining the end of the gene.