Mutants of Sindbis virus: I. Isolation and partial characterization of 89 new temperature-sensitive mutants

More than 100 new temperature-sensitive mutants of Sindbis virus have been isolated, following mutagenesis with nitrous acid, N-methyl- N′-nitro- N-nitrosoguanidine, 5-azacytidine, and 5-fluorouridine. Thirty-six of these mutants synthesize at least 60% as much RNA at the nonpermissive temperature as does the parental strain and are designated RNA +; 23 mutants synthesize between 10 and 60% as much RNA as the parental strain at 40° and are designated RNA ±; 30 mutants make less than 10% as much RNA at 40° and are called RNA −. The remaining mutants have not been tested for RNA incorporation. The thermal stability at 56° of most of the mutant particles has been examined. The majority of the RNA + mutants is more sensitive to heating at 56° than the parental HR strain, and RNA + mutations appear to reside primarily in genes coding for the structural proteins. Approximately 20% of either RNA ± or RNA − mutants are thermosensitive, and these mutations thus appear to reside primarily in genes coding for the nonstructural proteins. Complementation assays have been performed with a number of these mutants and with those of Burge and Pfefferkorn (1966a, b). The existence of three complementation groups among the RNA + mutants, which appear to encode the three major structural proteins, has been confirmed; no new complementation groups among RNA + mutants have been identified. A total of four complementation groups has been identified among the RNA − mutants. Thus, Sindbis virus contains at least seven complementation groups.