Inverted repeat preferences of Mos1 and Mboumar-9 transposases for in vitro DNA cleavage of transposon-containing plasmids.
A, schematic of the in vitro plasmid DNA cleavage assay and the expected products. The donor DNA plasmid contains a transposon comprising a kanamycin resistance gene (kanR) flanked by inverted repeats (black arrows), a chloramphenicol resistance gene (camR), and the oriR6K origin of replication. B, agarose gel of the products of in vitro IR DNA cleavage. Lane 1, 1-kb DNA ladder of markers (M); lane 2, pEPMosLL linearized with XbaI; lane 3, pEPMboLL digested with SacI to excise the transposon; lane 4, supercoiled (sc) plasmid. Lanes 5–7, cleavage of the Mos1 transposon containing two left IRs (LL), one left and one right IR (LR), or two right IRs (RR). Lanes 8–10, cleavage of Mboumar-9 transposons. C, quantification of the percentage of plasmid backbone released (as a proportion of the total intensity of the lane) in each of the reactions in lanes 5–10 above. The error bars indicate the S.D. between 4 independent measurements.