pT181 is a naturally occurring Staphylococcus aureus plasmid, encoding inducible resistance to tetracycline. The plasmid has a copy number of about 20 per cell, and belongs to the incompatibility group inc3. The complete nucleotide sequence of pT181 has been determined and consists of 4437 bp. The nucleotide sequence contains 69.8% A-T and 30.2% G-C pairs. pT181 was found to contain four open reading frames capable of coding for polypeptides containing more than 50 amino acids. All the putative polypeptides are coded by one strand. The molecular weights of the four putative polypeptides are (in daltons): A, 37,500; B, 35,000; C, 23,000, and D, 18,000. Polypeptide A corresponds to the repC protein, earlier shown to be specifically required for pT181 replication. Polypeptide B (and possibly polypeptide D) are involved in tetracycline resistance. No role has yet been established for polypeptide C; deletion of the coding sequence for the C polypeptide has no detectable effect on any property of the pT181 plasmid. A region consisting of about 1200 bp contains information for the replication and copy number control of this plasmid. The sequencing results are discussed in relation to the replication properties and tetracycline resistance associated with the pT181 plasmid.