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This study aimed to investigate the assembly characteristics of ciliature and cortical microtubules and the localization of tubulins in different depths and regions of the cortex. The hypotrichous ciliates have closely arranged cilia and a highly complex microtubular system. Direct fluorescence and immunofluorescence labeling were used to observe ciliary organelles and cortical microtubular cytoskeleton in Euplotes eurystomus. An immunofluorescence analysis demonstrated that α-tubulin localized to the ventral and dorsal ciliary organelles and their associated microtubules, while γ-tubulin localized to the basal bodies of ciliary organelles, macronuclear membrane, and excretory pore of a contractile vacuole in the interphase. A direct fluorescence analysis showed that the ciliature and cortical microtubules in the deep cortex were more clearly marked by fluorescent taxoid (FLUTAX). Interestingly, α- and γ-tubulins also colocalized to the ringlike ciliary base-associated microtubules of dorsal kineties. The short microtubular bundles between the bases of transverse cirri could be marked by FLUTAX and γ-tubulin rather than α-tubulin, suggesting that tubulins in cortical microtubules in E. eurystomus varied, and the differentiation of cortical microtubules in the hypotrichous ciliate tended to be versatile. Also, during morphogenesis, γ-tubulin also localized to the base of ciliary primordium, where new basal bodies were formed, suggesting that the endocellular position of γ-tubulin in ciliates was related to basal bodies and regulated by the cell cycle. This study might help understand the assembly characteristics and tubulin composition of microtubules in different depths and regions of the cortex in hypotrichous ciliate E. eurystomus. [ABSTRACT FROM AUTHOR]