We performed RNA sequencing on 18 total Western mosquitofish (Gambusia affinis) inhabiting one polluted and two unpolluted creeks in the Tar Creek Superfund site of northeastern Oklahoma, USA. Six female mosquitofish were sampled at each of the three sites for sequencing. After sequencing, we compared gene expression genome-wide between the polluted site and both unpolluted sites to identify genes with differential expression between polluted and unpolluted sites to help us uncover genes that may be involved in mediating persistence of mosquitofish in the toxic environment.