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Published on: February 15, 2012
TEMPERATURE-SENSTIVIE MUTANTS OF BACILLUS SUBTILIS BACTERIOPHAGE SP3. I. ISOLATION AND CHARACTERIZATION
Abstract:
Nishihara, Mutsuko (University of California, Los Angeles), and W. R. Romig. Temperature-sensitive mutants of Bacillus subtilis bacteriophage SP3. I. Isolation and characterization. J. Bacteriol. 88:1220-1229. 1964.-Twelve temperature-sensitive mutants of Bacillus subtilis bacteriophage SP3 were isolated from a suspension of the wild type treated with nitrous acid. The mutants were detected because of an impaired ability to replicate in the host cell at 45 C, in contrast to the parental wild type which has essentially the same burst size at temperatures between 37 and 48 C. However, the latent period of the wild-type phages was decreased as growth temperatures were varied from 37 to 49 C. The reduction of burst size at temperatures above 48 C is probably the result of the effect of higher temperatures on the metabolism of the host, because the bacterial growth rate is decreased between 45 and 51 C. A temperature effect on the mutants was studied by infecting cells at 45 C, and then transferring portions of the mixture to 37 C at two time intervals. One transfer was made within the expected 45 C latent period, and the other was made about 10 min after the end of the latent period. In both cases, removal of the phage-bacterium complexes from the inhibitory condition of high temperature did not result in an immediate increase of plaque-forming units. A delay was observed which might correspond to the time necessary to complete the temperature-sensitive synthesis in the infected cells. In experiments with three of the mutants, cells infected at 37 C were transferred to 45 C at various times during the 37 C latent period. The extent of inhibition upon transfer to the high temperature was found to decrease progressively with increasing time of prior incubation at 37 C.
Insights
Researchers isolated twelve temperature-sensitive mutants of Bacillus subtilis bacteriophage SP3. These mutants showed impaired replication at higher temperatures, indicating specific temperature-sensitive synthesis steps in phage development.
Area of Science:
- Microbiology
- Virology
- Bacteriophage Research
Background:
- Bacteriophage SP3 is a virus that infects Bacillus subtilis.
- Temperature-sensitive mutants can be valuable tools for studying essential gene functions.
- Understanding phage replication mechanisms is crucial in molecular biology.
Purpose of the Study:
- To isolate and characterize temperature-sensitive mutants of Bacillus subtilis bacteriophage SP3.
- To investigate the replication properties of these mutants at different temperatures.
- To elucidate the temperature-dependent stages of phage SP3 development.
Main Methods:
- Isolation of mutants using chemical mutagenesis (nitrous acid).
- Screening for temperature sensitivity based on plaque formation at restrictive temperatures (45°C).
- Growth curve analysis and temperature shift experiments.
Main Results:
- Twelve temperature-sensitive mutants of bacteriophage SP3 were successfully isolated.
- Mutants exhibited impaired replication at 45°C, unlike the wild-type phage.
- Temperature shift experiments revealed a delay in plaque formation, suggesting a requirement for completing temperature-sensitive synthesis.
- Incubation at a permissive temperature (37°C) prior to a restrictive temperature (45°C) reduced the inhibitory effect.
Conclusions:
- The isolated mutants are defective in essential phage functions that are temperature-sensitive.
- Phage SP3 development involves critical synthesis steps sensitive to temperature shifts.
- These mutants provide a valuable resource for dissecting the genetic and molecular basis of bacteriophage replication.
