Inbred mouse strains carrying spontaneous mutations in class I genes have been extremely informative in studies of the genetic mechanisms generating polymorphism in the major histocompatibility gene complex. In this report, we determine the molecular basis of the spontaneous loss mutation in BALB/c-H-2dm2 mice, which were previously shown not to express Ld antigens while maintaining normal expression of two other class I antigens, Kd and Dd. We show BALB/c-H-2dm2 mice do not transcribe detectable levels of Ld mRNA, indicating they do not produce a truncated Ld molecule as previously reported. Furthermore, in Southern blot comparisons using a series of low-copy genomic probes, the deletion was found to be approximately 140 kilobases and include the entire Ld gene along with three or more other class I genes mapping between Dd and Ld. These data represent direct genetic evidence for a spontaneous contraction in the genes encoding class I histocompatibility antigens, which in this case probably resulted from the misalignment of the 3' flanking regions of the Dd and Ld genes.