In earlier studies we identified in a human genomic library a gene (human relaxin gene H1) coding for a relaxin-related peptide. We now have evidence that the human genome possesses an additional relaxin-related gene (designated human relaxin gene H2) which appears to be selectively expressed in the ovary during pregnancy. Nucleotide sequence analysis revealed striking differences in the predicted structures of relaxin encoded by these two genes. Chemical synthesis of biologically active relaxin based on the sequence obtained from ovarian cDNA clones confirmed that the expressed gene (H2) encodes an authentic human relaxin. The expressed gene appears to be transcribed into two different sized mRNAs and preliminary evidence suggests that the mRNA transcripts possess different 3'-untranslated regions. There was no evidence for the expression of human relaxin gene H1 in the ovary and so far it is unclear whether gene H1 is expressed in another tissue or whether it represents a pseudogene. From the sequence data presented here it will now be possible to construct oligonucleotide probes and raise antibodies against synthetic peptides which could then be used to identify sites of relaxin biosynthesis and specifically quantitate the expression from either the H1 or H2 relaxin genes.