Slipped misalignment mechanisms of deletion formation: analysis of deletion endpoints

Abstract

To gain insight into the mechanisms of deletion formation between tandem repeats, Escherichia coli plasmids were engineered to carry a 101 bp tandem duplication within the tetA gene such that deletion of one of the repeats restores an intact tetA gene and tetracycline resistance to the cell. Four base-pair changes were introduced into one of the tandem repeats to serve as genetic markers. After selection for deletion, individual plasmid products were sequenced to deduce where within the repeat the deletion had occurred. Our analysis shows most deletions are fusions of the two repeats in a single 20 bp interval. This is consistent with the simple replication slip-pair model for deletion formation and suggests that this interval may have unusual features that promote deletion. Dimer replicon products have experienced a sister-chromosome exchange event in addition to deletion and carry two tetA loci: a deleted locus showing a similar distribution of endpoints as seen-in the monomer products and an unchanged repeat locus. Seemingly reciprocal dimers are occasionally recovered which carry both a deleted and a triplicated tetA locus. These are not truly reciprocal in that the sequence analysis showed that the deletion and triplication had occurred in separate intervals. Sequence analysis of the dimeric products is consistent with predictions from our sister-strand exchange model where slipped alignment of nascent DNA strands induces deletion formation concomitant with sister-chromosome exchange.