Large-scale selection and analysis of temperature-sensitive mutants for cell reproduction from BHK cells

Temperature-sensitive (ts)mutant cells for cell reproduction were isolated from the Syrian hamster cell line BHK21/13 by multiple culturing in the presence of 5-fluoro-2′-deoxyuridine (FdU) at 37.5° after N-methyl-N′-nitro-N-nitrosoguanidine mutagenesis. A simple method for cell fusion was devised, which enabled us to perform complementation studies with a large number of ts mutants. By using the method we have analyzed 219 ts mutants and classified them into 18 complementation groups. Mutants that belonged to the same complementation groups tended to exhibit similar patterns of inhibition of DNA synthesis at 39.5° however, some mutants belonging to the same group showed somewhat different patterns, probably due to occurrence of different mutations in the same gene. Distribution of the ts mutants among the 18 complementation groups was uneven; more than 50% of the mutants examined were assigned to complementation group B and G. The mutations belonging to complementation group B and G were found to be linked to the X chromosome.