Glyphosate-resistant (GR) Palmer amaranth (Amaranthus palmeri S. Watson) is considered one of the most troublesome weeds in the southern and central United States, but results of previous research to determine the mode of inheritance of this trait have been conflicting and inconclusive. In this study, we examined segregation patterns of EPSPS gene-copy numbers in F₁ and F₂ generations of A. palmeri and found no evidence of a Mendelian single-gene pattern of inheritance. Transgressive segregation for copy number was exhibited by several F₁ and all of the F₂ families, most likely the product of EPSPS copy-number variation within each plant. This variation was confirmed by assaying gene-copy number across clonal generations and among individual shoots on the same plant, demonstrating that EPSPS amplification levels vary significantly within a single plant. Increases and decreases in copy number occurred in a controlled, stress-free environment in the absence of glyphosate, indicating that EPSPS gene amplification is a random and variable process within the plant. The ability of A. palmeri to gain or lose EPSPS gene copies is a valuable adaptive trait, allowing this species to respond rapidly to selection pressures and changing environments.