Figure 6. The Cre–loxP system and its applications. (a) The loxP site (triangle) is a sequence of 34 bp composed of palindromic sequences of 13 bp separated by a sequence of 8 bp. Cre recombinase specifically recognizes this sequence, provokes the cleavage in DNA (vertical arrows) and (b) induces the recombination of DNA between the two loxP sites. This reaction is reversible. Several types of recombination events can be produced depending on whether the two loxP sites are carried by the same DNA molecule (recombination in cis) or by two different DNA molecules (recombination in trans) and depending on the respective orientation of the two loxP sites (the orientation of a loxP is given by the nonpalindromic 8-bp sequence). (c) Recombination in cis. If the two loxP sites have the same orientation, the DNA region situated between these sites is deleted during recombination. This type of configuration is used to create mutations devoid of the selection cassette (see Figure 5), deletions and conditional mutations (see Figure 7). If the orientation of the two loxP sites is opposed, recombination leads to the inversion of the region comprised between the two sites. (d) Recombination in trans. If one loxP site is integrated in the genome and the other is carried by a circular plasmid, there may be an insertion of sequences carried by the plasmid in the integrated loxP site. However, since the insertion is a rare event compared with deletion (i.e. the reverse reaction), this type of event requires the use of mutant loxP sites. When the loxP sites are both integrated in the genome, recombination in trans induces chromosomal rearrangements: deletions, duplications or translocations. Such recombination events are rare and have to be selected to be revealed. To do so, one can use truncated and nonfunctional hp-loxP and loxP-rt selection cassettes. After recombination between the loxP sites, and only in this case, a functional hp-loxP-rt cassette (the remaining loxP site is situated in an intron) is reconstituted, thus allowing selection of the chromosomal rearrangement desired (see legend to Figure 5). Furthermore, the relative orientation of loxP sites compared with the centromeric–telomeric axis of the chromosomes is important. Indeed, in the case of wrong relative orientation, recombination will result in the formation of acentric or dicentric chromosomes, which, in view of their great instability, will be eliminated and induce cell death.