Fig 5. Intptn3 inversion.
Plasmid pMC477 carries two copies of tRNALeu cloned in inverted orientation and separated by a 892bp spacer fragment (see Material and methods). The inverted repeats consist of the minimal tRNALeu 2–44 and the longer tRNALeu 2–88, both proficient in dimerization reactions. Plasmid pMC473 carries tRNALeu 2–44 and tRNAThr GQS_t10745, in inverted orientation as well. Both plasmids were incubated with IntpTN3 in a standard reaction (see Materials and methods). The NdeI restriction enzyme generates in each case two fragments of 2796 and 1777bp. Upon incubation with IntpTN3, NdeI digestion of pMC477 generates additional fragments of 2358 and 2215bp corresponding to the recombinant pMC477*. As for the integration/excision reactions, the relative intensity of the bands is compatible with an expected equilibrium reaction. We could not detect any inversion between tRNALeu and tRNAThr in plasmid pMC473.