TN5 IS50 TARGET RECOGNITION

This communication reports an analysis of Tn5/IS50 target site selecti on by using an extensive collection of Tn5 and IS50 insertions in two relatively small regions of DNA (less than 1 kb each). For both region s data were collected resulting from in vitro and in vivo transpositio n events. Since the data sets are consistent and transposase was the o nly protein present in vitro, this demonstrates that target selection is a property of only transposase, There appear to be two factors gove rning target selection. A target consensus sequence, which presumably reflects the target selection of individual pairs of Tn5/IS50 bound tr ansposase protomers, was deduced by analyzing all insertion sites. The consensus Tn5/IS50 target site is A-GNTYWRANC-T. However, we observed that independent insertion sites tend to form groups of closely locat ed insertions (clusters), and insertions very often were spaced in a 5 -bp periodic fashion. This suggests that Ta5/IS50 target selection is facilitated by more than two transposase protomers binding to the DNA, and, thus, for a site to be a good target, the overlapping neighborin g DNA should be a good target, too, Synthetic target sequences were de signed and used to test and confirm this model.