The murine Ly49 gene family is functionally analogous to the human killer cell Ig-like receptor (KIR) family of class I major histocompatibility complex (MHC) receptors. The number of KIR genes varies dramatically between individuals; however, the organization of the Ly49 genes has only been determined for the C57BL/6 (B6) mouse. The organization of the 129 Ly49 loci was determined from a BAC contig map by PCR and Southern blot analysis. In addition to the 10 Ly49 genes known from previous studies of the 129/J strain, 8 new genes were localized to the 129 Ly49 cluster. A gene order of Ly49q(1), e, (v, q(2)), e/c(2), l/r, s, t, e/c(1), r, u, u/i, i(1), g, p/d, (i(2), p), and o was determined. The 129 Ly49 gene cluster is predicted to span approximately 600 kb. These results indicate that Ly49 gene numbers can be significantly different between inbred mouse strains, analogous to the haplotype differences observed in the human KIR genes.