Figure 6.

DNA topology of recombination products. Recombination of circular substrates containing pairs of core site variants in the absence (panels I–III) or presence (panels IV–VII) of directly repeated DR1–DR2 accessory motifs were treated with TnpI or left untreated as indicated. Reactions were singly nicked with DNase I and run on a 0.8% agarose gel. Positions of the open circular (ocS), linear (lin), and supercoiled (scS) forms of the substrates are indicated, as are the positions of the nicked two-, three-, four-, five-, six-, seven- and eight-noded catenane and knot products of recombination. Note that the majority of secondary (twist) knots generated by consecutive rounds of recombination at Core1.2 (panel I) migrate slightly ahead of the primary (torus) knots arising from recombination at the symmetrical Core1.1 and Core2.2 sites (panels II and III). Recombination of the DR1 and DR2-containg plasmids (panels IV–VII) yielded two-noded catenanes (ocCat-2) as the only product of recombination. Additional bands (asterisk) correspond to circular deletion products that were released from catenanes by over digestion with DNase I.