Figure 5.

TnpI-mediated recombination at symmetrical core sites in vitro. Reporter plasmids carrying tandem copies of the wild-type (Core1.2) or symmetrical (Core1.1 and Core2.2) core sites under the control of DR1–DR2 accessory sequences (panels IV–VI) or not (panels I–III) were incubated with purified TnpI for the indicated times. Reactions were digested with SmaI (SI) and analysed by agarose gel electrophoresis. The cartoon summarizes all possible DNA fragments that may arise from recombination at both substrate sites (site1 and 2) after digestion with SmaI. S1 and S2 are the initial substrate fragments. D1 and D2 are DNA fragments arising from recombination between directly oriented sites1 and 2. Inv1 and Inv2 are recombination products of inversely oriented sites. Inter1, Inter2, Inter3 and Inter4 are specific ‘intermolecular’ site1 × site1 and site2 × site2 recombination products in which flanking DNA segments are joined in head-to-head and tail-to-tail configuration (Inter4 was too small to be seen on the gel). HJ intermediates migrate as a χ-form at an upper position of the gel.