DNA sequence analysis of spontaneous tonB deletion mutations in a polA1 strain of Escherichia coli K12.

By DNA sequence analysis, we have determined a spectrum of 61 spontaneous mutations occurring in the endogenous tonB gene in the polA1 strain of Escherichia coli. The overall mutation frequency was approximately 2.4-fold higher in the polA1 strain and this was attributable to enhanced rates of deletion and frameshift mutations. Among 39 deletions, a hot spot (17 mutations) was detected: a 13-bp deletion presumably directed by a 3-bp repeated sequence at its end points. The remaining 22 were distributed among 19 different mutations either flanked (16/19) or not flanked (3/19) by repeated sequences. Single-base frameshifts accounted for 8 mutations of either repeated (3/8) or nonrepeated (5/8) bases among which 6 were minus one frameshift. In contrast to previous reports, we did not frequently observe a 5'-GTGG-3' sequence in the vicinity of the deletions and frameshifts. The results presented here indicated an anti-deletion and anti-frameshift role for DNA polymerase I.

Asymmetric crossing over in the spontaneous formation of large deletions in the tonB-trp region of the Escherichia coli K-12 chromosome.

The chromosomal tonB gene of Escherichia coli was used as a target for the detection of spontaneous deletion mutations. The deletions were isolated in both recA+ and recA- cells, and mutants carrying large deletions were identified because they also lacked part or all of the trp operon. The frequencies of tonB-trp deletion were 1.79 x 10(-9) and 1.09 x 10(-9) for recA+ and recA- cells, respectively. We analyzed 12 deletions from recA+ and 10 from recA- cells by cloning and direct sequencing. The deletions ranged in size from 5612 bp to 15142 bp for recA+ and from 5428 bp to 13289 for recA- cells. Three deletions from recA+ cells and five deletions from recA- cells were found to have occurred between short sequence repeats at the termini of the deletion, leaving one copy of the repeat in the mutant sequence. Seven deletions from recA+ cells and three deletions from recA- cells did not have repeats at their termini; in these cases, the DNA sequences that are adjacent to the deletion termini in the wild-type are characterized by short (2-4 bp) repeats. From these results, a model is presented for the generation of deletion mutations which involves formation of an asymmetric crossover mediated by repeated sequences of 2- to 4-bp.