Answer T2

From 2006.igem.org

1. From what we have seen, once an invertasome is formed with two hix sites, recombination can occur multiple times.
2. A new invertasome would have to form with new hix sites to cause a new recombination.
3. Each time recombination occurs, two negative supercoils are used, a topological knot is produced, and the internal halves of two hix sites are exchanged.