Plasma kallikrein (Fletcher factor) is a hepatic serine proteinase that participated in the early phase of blood coagulation. From two genomic libraries, we succeeded to isolate four overlapping clones representing the entire rat plasma kallikrein gene. Using selective DNA sequencing, polymerase chain reactions, and restriction mapping, we demonstrated that the gene for rat plasma kallikrein was 22 kb in length. Similar to human factor XI [Asakai et al. (1987) Biochemistry 26, 7221-7228], we also found that the plasma kallikrein gene is composed of 15 exons and 14 introns. A potential transcription initiation step was determined by a novel application of the polymerase chain reaction technique. Computer analysis of the 5'-promoter region of this gene revealed some putative control elements that might regulate the rat plasma kallikrein gene expression. These data and the results of chromosomal localization reported in the present study for mouse (chromosome 8) and human (chromosome 4) plasma kallikrein genes strongly corroborate a genic duplication event from a common ancestor to both plasma kallikren and factor XI.