After accumulating mutations by the aid of marked inversions, spontaneous occurrence rates of chromosome aberrations were estimated for 1148 chromosome lines that originated from five stem line second chromosomes of Drosophila melanogaster. In chromosome lines originating from three stem chromosomes (CH, PQ, and RT), mutations were accumulated for 7550, 7252, and 7256 chromosome generations, respectively, but no structural change was detected. For the chromosome lines that originated from the other two stem chromosomes, the situation was different: Twenty aberrations (19 paracentric inversions and 1 translocation between the second and the third chromosomes) during 45990 chromosome generations took place in the 500 chromosome lines derived from stem line chromosome (AW), and 92 aberrations (83 paracentric inversions, 6 pericentric inversions, 2 translocations between the second and the third chromosomes and 1 transposition) arose during 45006 chromosome generations in the 500 chromosome lines derived from stem line chromosome (JH). For the AW group the occurrence rate becomes 0.00043 per chromosome per generation for all aberrations and 0.00041 for inversions. For the JH group the corresponding rates are 0.00204 and 0.00198, respectively.—A non-random distribution of the breakpoint on the salivary gland chromosome was observed and the breakpoints were concentrated in the regions 26, 29, 33, and 34.—The cytoplasms and the chromosomes (other than the second chromosomes) were made approximately uniform throughout the experiments. Thus, this remarkable variability in the occurrence rate is most probably due to the differences in one or more chromosomal elements on the original five stem chromosomes. The mutable chromosomes (AW and JH) appear to carry a kind of mutator factor such as hi (Ives 1950).