The amino acid analog albizziin was used to isolate Chinese hamster ovary cell lines which overproduce asparagine synthetase. Mutants selected in a single step after ethyl methane sulfonate mutagenesis were approximately 10-fold more resistant to the drug than the parental lines and expressed 8- to 17-fold elevations in enzyme activity. The karyotypes of these lines show alterations such as breaks and translocations affecting the long arm of chromosome 1. Cell lines isolated in several steps by growth in progressively increasing concentrations of albizziin were more resistant to the drug and exhibited up to 300-fold enhancement of asparagine synthetase activity. The multistep albizziin-resistant cell lines usually had expanded chromosomal regions which stained somewhat homogeneously, often on the long arm of chromosome 1. These results suggest that resistance to albizziin in the multistep lines may be due to gene amplification.