Two cDNA clones for complement component C2 have been isolated from a high-complexity human liver cDNA library by using a mixture of 64 synthetic oligonucleotides as a probe. The 400-base-pair insert of pC201 codes for a region containing the active site serine residue and the secondary substrate binding pocket of the serine protease. This part of C2 is 34% homologous to the corresponding region of the related serine protease factor B and additional similarity is evident from a number of conservative amino acid replacements in this region. The insert of pC201 was used as a specific probe in RNA transfer analysis to determine the size of the C2 mRNA as approximately equal to 2.9 kilobases. Southern blot analysis of genomic DNA of unrelated individuals identified a single C2 locus and showed no cross-hybridization with the factor B locus.