Physical mapping of DNA clones by nonisotopic in situ hybridization has greatly facilitated the human genome mapping effort. Here we combine a variety of in situ hybridization techniques that make the physical mapping of DNA clones to mouse chromosomes much easier. Hybridization of probes containing the mouse long interspersed repetitive element to metaphase chromosomes produces a Giemsa-like banding pattern which can be used to identify individual Mus musculus, Mus spretus, and Mus castaneus chromosomes. The DNA binding fluorophore, DAPI, gives quinacrine-like bands that can complement the hybridization banding data. Simultaneous hybridization of a differentially labeled clone of interest with the banding probe allows the assignment of a mouse clone to a specific cytogenetic band. These methods were validated by first mapping four known genes, Cpa, Ly-2, Cck, and Igh-6, on banded chromosomes. Twenty-seven additional clones, including twenty anonymous cosmids, were then mapped in a similar fashion. Known marker clones and fractional length measurements can also provide information about chromosome assignment and clone order without the necessity of recognizing banding patterns. Clones hybridizing to each murine chromosome have been identified, thus providing a panel of marker probes to assist in chromosome identification.