Mutations of temperature sensitivity in R plasmid pSC101

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Abstract
Temperature-sensitive (Ts) mutant plasmids isolated from tetracycline resistance R plasmid pSC101 were investigated for their segregation kinetics and DNA replication. Apparently, multiple copies of a plasmid are distributed to daughter cells [Escherichia coli] in a random fashion and are thus diluted out when a new round of plasmid DNA replication is blocked. When cells harboring type I mutant plasmids were grown at 43.degree. C in the absence of tetracycline, antibiotic-sensitive cells were segregated after a certain lag time. This lag most likely corresponds to a dilution of plasmids existing prior to the temperature shift. The synthesis of plasmid DNA in cells harboring type I mutant plasmids was almost completely blocked at 43.degree. C. These plasmids may have mutations in the gene(s) necessary for plasmid DNA replication. Cells harboring a type II mutant plasmid exhibited neither segregation due to antibiotic sensitivity nor inhibition of plasmid DNA replication throughout cultivation at high temperature. The type II mutant plasmid probably has a temperature-sensitive mutation in the tetracycline resistance gene. Antibiotic-sensitive cells harboring type III mutant plasmids appeared at high frequency after a certain lag time, and the plasmid DNA synthesis was partially suppressed at the nonpermissive temperature. They exhibited a pleiotropic phenotype, such as an increase of drug resistance level at 30.degree. C and a decrease in the number of plasmid genomes in a cell.