A mouse cDNA probe homologous to the human MCF2 transforming sequence has been identified and partially cloned, and is used here to localize the gene on the mouse X chromosome. The human gene has been physically mapped to within 60 kb of the gene for coagulation factor IX, within a large conserved linkage group between the mouse and human genomes which extends from HPRT to G6PD on the X chromosomes of both mammalian species. In situ hybridization of the mouse Mcf-2 probe onto mouse metaphase chromosomes indicates that this gene lies in the same region of the X chromosome as Cf-9, the mouse gene for coagulation factor IX. Moreover, segregation of species-specific genomic DNA polymorphisms for Mcf-2 and Cf-9 in a total of 203 individuals derived from two large interspecific mouse backcross populations (which are also segregating for 17 other X-linked molecular markers) demonstrates that the mouse genes are separated by only 0.5 +/- 0.5 cM. Despite this short distance we were able to order Mcf-2 and Cf-9 relative to one another and other genes in this region. The mouse gene order Hprt-Cf-9-Mcf-2-G6pd predicts a similar ordering of genes on the human X chromosome, a gene order which has only recently been demonstrated by physical mapping. Thus, the map location and linkage relationships of the Mcf-2 gene are similar in man and mouse, and this unique protooncogenic locus is part of a conserved linkage group on the mammalian X chromosome.