The pericentromeric region of the human X chromosome is characterized by a tandemly repeated family of 2.0 kilobasepair (kb) DNA fragments, initially revealed by cleavage of human DNA with the restriction enzyme BamHI. We report here the complete nucleotide sequence of a cloned member of the repeat family and establish that this X-linked DNA family consists entirely of α satellite DNA. Our data indicate that the 2.0 kb repeat consists of twelve α satellite monomers arranged in imperfect, direct repeats. Each of the αX monomers is approximately 171 basepairs (bp) in length and is 60–75% identical in sequence to previously described primate a satellite DNAs. The twelve αX monomers are 65–85% identical in sequence to each other and are organized as two adjacent, related blocks of five monomers, plus an additional two monomers also related to monomers within the pentamer blocks. Partial nucleotide sequence of a second, independent copy of the 2.0 kb BamHI fragment established that the 2.0 kb repeat is, in fact, the unit of amplification on the X. Comparison of the sequences of the twelve αX monomers allowed derivation of a 171 bp consensus sequence for a satellite DNA on the human X chromosome. These sequence data, combined with the results of filter hybridization experiments of total human DNA and X chromosome DNA, using subregions within the 2.0 kb repeat as probes, provide strong support for the hypothesis that individual human chromosomes are characterized by different α satellite families, defined both by restriction enzyme periodicity and by chromosome-specific primary sequence.