The Tn10 tetR gene encodes the repressor that regulates transcription of the Tn10 tetracycline resistance determinant. We have determined the DNA sequence of the tetR gene and a 905 base pair region immediately 3' to tetR. The tetR gene is located on a 701 base pair HincII restriction fragment. Deletions at either end of this region eliminate synthesis of the wild-type TetR protein in E. coli minicells, and eliminate TetR activity as measured by repression of beta-galactosidase synthesis in tetA-lacZ operon fusion strains. Taken together, the DNA sequence and the genetic data indicate that tetR encodes a 207 amino acid protein with a calculated molecular weight of 23,328. This value is in good agreement with estimates of 23,000-25,000 based on electrophoretic mobility in SDS-polyacrylamide gels. There is 47% amino acid sequence homology between the deduced sequences of the Tn10 and RP1/Tn1721 TetR proteins. There is, in addition, significant amino acid sequence homology between an NH2-terminal region of the Tn10 TetR repressor and the DNA recognition regions of other DNA-binding proteins.