In order to ascertain that DNA methylation is associated with the formation of sister chromatid exchange (SCE), we investigated the SCE frequency on allocyclic replicating X chromosomes induced by 5-Azacytidine (5-AzaC) using lymphocytes from human peripheral bloods. Autoradiography was used for discrimination. between early replicating S (X^a) which is genetically active and late replicating X (X^(in)) and acridine orange and Q-banding techniques for sister chromatid differential staining and identification of individual chromosomes, respectively. These three methods were performed on the same metaphase. The SCE frequency of X^(in) in lymphocytes was found to be slightly high compared with that of X^a when exposed to 5-AzaC (X^(in)/X^a = 1.38) whereas the SCE frequency of X^(in) showed the two-fold increase compared with that of X^a when unexposed (X^(in)/X^a = 2.00). These results proved statistically significant reflecting that 5-AzaC actually acts as an inhibitor of SCE induction only in X^(in)(p<0.01) although the SCE frequency in 5-AzaC-exposed cultures was higher than that in unexposed ones. Interestingly, it was found that the SCE occurring sites in X^(in) were concentrated on the Xq21 which known to be the inactivation center of X chromosome (36% of total SCE of X^(in)). These findings suggest that hypo(de)methylation induced by 5-AzaC in methylated X^(in), especially in the highly methylated inactivation center, might induce some alteration in the process of DNA replication and/or repair, which are responsible for the SCE formation.