The IS Dimer Circular Intermediate Participates in Transposition.

The mobile colistin resistance gene is a serious threat to global human and animal health. The composite transposon Tn and its circular intermediate were proposed to be involved in the spread of but their roles remain poorly understood. To further explore the intermediates during the transposition of Tn, we engineered strains that carry an intact Tn transposon or its deletion derivatives. PCR assays were performed to detect IR-IR junctions and possible circular intermediates. We carried out transposition experiments to calculate transposition frequency. The transposition sites were characterized by whole genome sequence and ISMapper-based analyses. The presence of an intact Tn was demonstrated to be essential for the successful transposition of , although both Tn and Tn-ΔIR could form circular intermediates. The insertion sequence junction structure was observed in all constructed plasmids but the IS dimer was only formed in one construct containing an intact Tn. The average frequency of transposition in an strain possessing an intact Tn was ∼10 per transformed cell. We identified 27 integration sites for the Tn transposition event. All the transposition sites were flanked by 2 bp target duplications and preferentially occurred in AT-rich regions. These results indicate that transposition relies on the presence of an intact Tn. In addition, formation of the tandem repeat IS could represent a crucial intermediate. Taken together, the current investigations provide mechanistic insights in the transposition of .