Most mouse-human somatic cell hybrids result in preferential loss of human chromosomes, absence of human 28S ribosomal RNA, and suppression of human nucleolus organizer. DNA methylation has been known to be closely related with the inhibition of ribosomal RNA synthesis in human cells. On the basis os these aspects, this experiment has been focused on whether there is a direct relationship between chromosome segregation and DNA methylation in hybrid condition or not. The hybrids were obtained from fusion between HE2301, a human embryo fibroblast and A9, a mouse L fibroblast. Cloning of these hybrids was performed using limiting dilution, and 4 clones obtained through this process were used. These clones were exposed to 5-azacytidine (5-AzaC) which inhibits methylation of newly synthesized DNA. It was found that there is no difference between the number of active NOR in A9 exposed to 5-AzaC and in unexposed the 4 clones. However, the mean number of active NOR in 4 clones exposed to 5-AzaC was found to be significantly increased. These results may imply that net increase of the activated NOR results from reactivation of the inactivated human NORs by 5-AzaC. Thus, the reason for the reactivation of human NORs seems to be induced by hypo- or demethylation of human DNA. I suggest that the segregation of human chromosomes in mouse-human hybrid may be directly associated with the degree of methylation in human NORs.