We have cloned the UNI3 gene in Chlamydomonas and find that it encodes a new member of the tubulin superfamily. Although Uni3p shares significant sequence identity with α-, β-, and γ-tubulins, there is a region of Uni3p that has no similarity to tubulins or other known proteins. Mutant uni3-1 cells assemble zero, one, or two flagella. Pedigree analysis suggests that flagellar number in uni3-1 cells is a function of the age of the cell. The uniflagellate uni3-1 cells show a positional phenotype; the basal body opposite the eyespot templates the single flagellum. A percentage of uni3-1 cells also fail to orient the cleavage furrow properly, and basal bodies have been implicated in the placement of cleavage furrows in Chlamydomonas. Finally when uni3-1 cells are observed by electron microscopy, doublet rather than triplet microtubules are observed at the proximal end of the basal bodies. We propose that the Uni3 tubulin is involved in both the function and cell cycle-dependent maturation of basal bodies/centrioles.