cDNA encoding the p70 polypeptide subunit of the human Ku autoantigen was isolated. In vitro expression analysis of the cDNA demonstrates that it encodes the entire open reading frame. Nucleotide sequence analysis and comparison to other previously described sequences indicate the existence of several single-nucleotide and amino acid polymorphisms. Southern blot analyses demonstrate the presence of multiple copies of homologous DNA sequences in the human genome. These data support the hypothesis that multiple genes encode a family of Ku(p70)-related polypeptides.