1,3-Butadiene (BD) is carcinogenic and mutagenic in B6C3F1 mice. We determined the lacI mutant frequency and mutational spectrum in spleen following inhalation exposure to BD at levels that are known to induce tumors. B6C3F1 lacI transgenic mice were exposed to air or to 62.5, 625, or 1250 ppm BD for 4 weeks (6 h/day, 5 days/week) and euthanized 14 days after the last exposure. BD increased the lacI mutant frequency in spleen at all levels of BD examined. In BD-exposed mice, an increased frequency of G:C-->A:T transitions occurred at non-5'-CpG-3' sites. Exposure to BD in B6C3F1 lacI transgenic mice also increased the frequency of base substitution mutations that occurred at A:T base pairs when compared to air controls. The increased frequency of specific mutations at G:C base pairs in spleen was not observed in our previous studies in bone marrow and indicates tissue-specific differences in the BD-induced mutational spectrum. These data demonstrate that in vivo transgenic mouse mutagenicity assays can identify tissue-specific mutagenicity and mutational spectrum responses of genotoxic carcinogens at exposure levels that are known to induce tumors.