Many selectable mutants of somatic Chinese hamster cells have been described, but very few of the mutations have been mapped to specific chromosomes. We have utilized the microcell-mediated gene transfer technique to establish the location of three selectable genetic markers on chromosome 2 of Chinese hamster. Microcells were prepared from the methotrexate-resistant MtxRIII line of Flintoff et al. (Somatic Cell Genet. 2:245-261, 1976) and fused to wild-type CHO cells, and microcell hybrids (transferants) were selected in medium containing methotrexate. All transferants were karyotyped and found to contain a marker chromosome from the donor MtxRIII line. This marker chromosome, called 2p-, consisted of a chromosome 2 with a reduced short arm resulting from a reciprocal translocation between 2p and 5q. In experiments utilizing emetine-resistant (Emtr) or chromate-resistant (Chrr) recipient cells it was found that the emt+ and chr+ wild-type genes were cotransferred with the 2p- chromosomes. Karyotype analysis of several transferants with rearranged or broken 2p- markers allowed regional localization of the emt and chr loci to the proximal third of the long arm and localization of the gene or genes conferring methotrexate resistance to the short arm. These results confirm our earlier assignment of the emt and chr loci to chromosome 2 in Chinese hamster.