Metabolic resistances to atrazine (atz-R) and mesotrione (meso-R) occur in several waterhemp [Amaranthus tuberculatus (Moq.) Sauer] populations in the United States. Interestingly, although metabolic atz-R but mesotrione-sensitive A. tuberculatus populations have been reported, an Amaranthus population has not been confirmed as meso-R but atrazine-sensitive, implying an association between these traits. Experiments were designed to investigate whether the single gene conferring metabolic atz-R plays a role in meso-R. An F₂ population was generated from a multiple herbicide–resistant A. tuberculatus population from McLean County, IL (MCR). A cross was made between a known meso-R male clone (MCR-6) and a herbicide-sensitive female clone from Wayne County, IL (WCS-2) to develop an F₁ population. Survival of MCR-6 plants following atrazine POST treatment (14.4 kg ha^–1) indicated the male parent was homozygous atz-R. F₁ plants were intermated to obtain a segregating pseudo-F₂ population. Dose–response and metabolic studies conducted with mesotrione using F₁ plants indicated intermediate biomass reductions and metabolic rates compared with MCR-6 and WCS. F₂ plants were initially treated with either mesotrione (260 g ha^–1) or atrazine (2 kg ha^–1) POST, and after 21 d of recovery, vegetative clones from surviving resistant plants were subsequently treated with the other herbicide. When mesotrione was applied first, the meso-R frequency was 8.2%, and when atrazine was applied first, the atz-R frequency was 75%. However, the meso-R frequency increased to 16.5% following preselection for atz-R, and 100% of surviving meso-R plants were atz-R. Our findings indicate that the gene conferring metabolic atz-R is also involved with the meso-R trait within the population tested.