Tandem duplication induced by an unusual ampA1-, ampC-transducing lambda phage: a probe to initiate gene amplification

Abstract

Secondary attachment site lambda-lysogens were isolated in an Escherichia coli strain carrying multiple tandem 9.8 kb repeats. The repeat carried the structural gene for chromosomal beta-lactamase, ampC. One lysogen produced lysates with amp-transducing activity. Three types of phages with different densities were obtained from this lysogen. The one with the lowest density was found to be a helper lambda cI857S7 phage. The other two phage showed identical restriction endonuclease fragmentation patterns. The difference in density was due to the presence or absence of phage tail. In lambda damp the right cohesive end segment was deleted in a random fashion with the majority ending between 81.0% and 82.4% of lambda. The chromosomal segment of lambda damp was most likely located at the lambda attachment site. The lambda damp DNA was compared to that of ColE1 hybrid carrying the chromosomal amp segment and a ColE1 hybrid carrying the same 9.8 kb amp repeat as the lysogen from which lambda damp was isolated. It was found that the chromosomal part of lambda damp constituted 9.8 kb, i.e. the size of one repeat. Moreover, the novel joint between adjacent repeats was present. In a lambda attB-deleted E. coli K-12 strain, lysogenic for lambda damp, highly ampicillin-resistant mutants occurred at an exceedingly high frequency. They were found to contain in the chromosome an amplified 9.8 kb repeat. This suggested that integration of the novel joint for lambda damp into the amp region gives rise to an amplifiable duplication. In E. coli lysogenized for lambda damp at lambda attB highly ampicillin-resistant clones were also found at a high frequency. These clones carried multiple tandem repeats of lambda damp DNA, each with an intact right end segment.