Southern blot analysis with a murine interferon-alpha2 (MuIFN-alpha2) cDNA probe revealed restriction fragment polymorphism of EcoRI- and HindIII-digested C57BL/6 and BALB/cDNA. The inheritance pattern of this polymorphism was examined using DNA from each of the seven recombinant inbred strains derived from C57BL/6 and BALB/c; the strain distribution pattern suggests linkage of INF-alpha genes to two histocompatibility loci on chromosome 4. Southern blot analysis of DNA from six bilinear congenic strains carrying different fragments of the BALB/c chromosome 4 on a C57BL/6 background showed linkage of IFN-alpha genes to the histocompatibility locus H-15. It can therefore be concluded that the IFN-alpha gene cluster is situated on chromosome 4 near the H-15 locus, between loci Mup-1 and b.