Most stationary-phase Lac+ revertants do not display heritable stationary-phase mutator phenotypes

Frameshift-bearing strainExpt.Cumulative Lac+ colonies on day 5 per 108 viable cellsaReversion relative to Tn-less parentReversion relative to Tn-carrying control strainPhenotype
Tn-less parent
FC40197.9 ± 111
236.1 ± 8.21
321.5 ± 101
Tn10-containing control strain
SMR37391158 ± 131.6
2110 ± 143
362.1 ± 122.9
Recycled Lac strains
SMR376819.1 ± 1.30.06antimutator
SMR37691132 ± 100.8nonmutator
SMR37701279 ± 351.8
338 ± 30.6nonmutator
SMR37711238 ± 311.5nonmutator
SMR3772114.3 ± 2.70.09
35.2 ± 1.50.08antimutator
SMR37732200 ± 641.8
339.3 ± 4.10.6nonmutator
SMR3774278.7 ± 110.7nonmutator
SMR3775261.5 ± 6.60.6nonmutator
SMR3776262.3 ± 6.30.6nonmutator
SMR3777243.6 ± 3.90.4nonmutator
SMR3778254.7 ± 7.40.5nonmutator
SMR3779273.4 ± 4.80.7nonmutator
Nonmutator mean ±SE0.83 ± 0.1
  • Twelve independent recombination-dependent stationary-phase Lac+ revertants were “recycled” back to carrying their original lac + 1 frameshift mutation by transduction with P1 grown on zah-281::Tn10 lacI33-carrying strain SMR3739 (this work). SMR3739 was made by transduction of zah-281::Tn10, a transposon linked with lac, from strain CAG12080 (Singer et al. 1989) into the lac + 1 frameshift-bearing strain FC40 (Cairns and Foster 1991). By this means, each recycled Lac strain (SMR3768-3779; this work) carries the original lac + 1 frameshift allele and now also carries the transposon zah-281::Tn10 linked with lac. (The Tn was used as a selectable marker in these transductions.) These recycled Lac strains also carry any unselected mutations that may have occurred in association with recombination-dependent hypermutation (Torkelson et al. 1997) and are tested here for whether or not they possess a heritable mutator phenotype for stationary-phase Lac+ reversion as follows:

  • Recycled Lac strains were subjected to prolonged incubation on minimal medium with lactose as the sole carbon source, and Lac+ stationary-phase mutants were scored over five days’ incubation as described previously (e.g., Harris et al. 1994, 1996). For each strain in each experiment (Expt.), 8-12 independent cultures were used and the mean frequency of Lac+ colonies ±1 SE is given. The presence of zah-281::Tn10on the F′ appears to cause a roughly twofold increase in stationary-phase Lac+ reversion relative to the Tn-less parent. This might be caused by the transposon insertion itself or by transposon hopping or excision in stationary phase that could provide an additional source of DNA double-strand breaks, which are necessary for recombination-dependent stationary-phase mutation (Harris et al. 1994; reviewed by Rosenberg 1997). The recycled strains that have been through one round of recombination-dependent stationary-phase mutation, in addition to carrying zah-281::Tn10, do not display obvious increases in stationary-phase mutability relative to the Tn-carrying control strain. In addition, two recycled strains seem to have accrued mutations that inhibit their ability to engage in recombination-dependent Lac reversion and are designated antimutators. These two strains are not merely slow colony formers (data not shown) but appear to be genuine stationary-phase mutation-impaired strains.

  • a Values are mean ± SEM.