Stable mutants highly resistant to the protein-synthesis-inhibitor diphtheria toxin have been selected in Chinese hamster ovary (CHO) cells. Protein synthesis in extracts of mutant cells is resistant to the inhibitory action of diphtheria toxin, indicating that the lesion has affected the protein-synthesis machinery. However, about 50% of the elongation factor-2 (EF-2) activity in the mutant cells can still be ADP-ribosylated by diphtheria toxin, and this remaining EF-2 activity is similar to that present in the wild-type cells. We suggest that this result is best explained by assuming that our CHO cells contain two functional copies of the EF-2 gene, and that only one of the copies is altered in the mutants. According to this view, the mutated allele produces EF-2 resistant to ADP-ribosylation which is capable of supporting cell growth in the presence of diphtheria toxin. Although the Dipr marker seems to act dominantly in the parental CHO cells, its behavior in Dipr X Dips hybrids (CHO X CHO) is recessive as measured by cell survival in presence of the toxin. This paradoxical behavior may be due to a gene dosage effect. Segregation studies from hybrids show that the Dipr marker segregates independently of the Emtr and Thgr markers indicating that the Dipr locus is not linked to either the Emtr locus or to the X chromosome.