To better characterize abnormalities affecting rat chromosome 1 during mammary carcinogenesis, tumors were induced by nitrosomethylurea in F1 hybrid rats polymorphic at multiple chromosome 1 loci. By means of restriction fragment length polymorphism and microsatellite length polymorphism analyses, we observed loss of heterozygosity or allelic imbalance affecting various loci on the q arm of chromosome 1 in a high percentage of the 49 tumors analyzed. Fifty percent of the tumors showed loss or imbalance affecting the most distal (1q55) INS1 (rat insulin 1 gene) locus. The MT1PA (metallothionein-1 pseudogene a) locus was observed to be affected in 58% of tumors induced in BUF/NCr x ACI/Vsp rats. Most of the losses appeared to have occurred by mitotic recombination. No parental bias was observed on the affected chromosome 1. Tumors were also screened for mutations in codon 12 of the Ha-ras-1 gene, which is located on 1q. We observed an association between the presence of mutation and allelic imbalance. These studies confirm our previous cytogenetic observations of a high level of nonrandom instability affecting rat chromosome 1 during mammary carcinogenesis. The observed loss of heterozygosity may indicate the existence of a putative tumor suppressor gene within the distal half of the 1q arm. These abnormalities, however, could also be related to the early stages of Ha-ras amplification.