Targeted gene disruption was used to investigate the function of MYO1, an unconventional myosin gene in Tetrahymena thermophila. Phenotypic analysis of a transformed strain that lacked a functional MYO1 gene was conducted at both 20 °C and 35 °C. At either temperature the Δ MYO1 strain had a smaller cytoplasm/nucleus ratio than wild type. At 20 °C, Δ MYO1 populations had a longer doubling time than wild type, lower saturation density, and a reduced rate of food vacuole formation. However, at 35 °C, these characteristics were comparable to wild type. Although micronuclear division and cytokinesis appeared normal in Δ MYO1 cells, failure of the macronucleus to elongate properly resulted in unequal segregation of macronuclear DNA in cells maintained at either 20 °C or 35 °C.