BALB/c 3T3 cells transformed in vitro with a temperature-sensitive mutant of polyoma virus were cloned. Forty-eight clones examined demonstrated heterogeneity with respect to doubling-time in vitro and tumorigenicity in syngeneic mice in vivo. Observation periods that lasted in certain cases as long as 2 years showed that some clones exhibited a relatively high tumorigenicity, i.e., they yielded a relatively high incidence of tumors following a small inoculum of cells and a relatively short latency period. Other clones were relatively low tumorigenic: even high tumor cell inocula yielded a relatively low tumor incidence following a relatively long latency period. These results indicate that at least in this system variation in tumorigenicity is generated independently of host factors. An intraclonal heterogeneity with respect to the length of the precancer latency period was seen. Some tumors appeared relatively early following inoculation of cloned cells, whereas others appeared considerably later following an identical inoculum of the same clone. Cloned in vitro transformed cells were passaged once in syngeneic mice and recultured. The single in vivo passage cycle augmented considerably the tumorigenicity of these cells as compared to their in vitro maintained clonal ancestors. The increased tumorigenicity of the in vivo passaged cells is due, most probably, to the in vivo induction and/or selection of high tumorigenic intraclonal variants. The survival time of mice bearing high tumorigenicity variants was very similar to that of mice bearing low tumorigenicity variants.