Chromosome preparation in late metaphase cells from bone marrow of colcemid treated male Chinese hamsters were used to analyse the sequence of separation of sister centromeres. Chromatids of chromosomes 2 and 1 are the first ones to separate at centromeres, followed by members of group B, D and C. Some acrocentric chromosome is always the last one to separate at the centromere. The data point to a possible correlation between the position of a centromere in the separation sequence in the genome and the amount of centromeric heterochromatin as well as relation to the phenomenon of non-disjunction.