[1P-78] An improved chemically-inducible Cre-loxP recombination system with no leakiness
Site-specific DNA recombination systems represented by Cre-loxP are a powerful and useful tool for modifying transgenes in chromosomes of living organisms. A variety of chemically-inducible Cre-loxP recombination systems have been developed, allowing for conducting conditional genome engineering. However, conventional systems still suffer from the leakiness of DNA recombination in the absence of its chemical compound. To overcome this limitation, here we report an improved destabilized Cre recombinase (DD-Cre 2.0) that is based on the multiple tandemly-repeated destabilizing domains (DD) derived from E. coli DHFR to enhance its degradation by the proteasomal pathway. In the absence of Trimethoprim (TMP) which is a protein-stabilizing ligand for ecDHFR, DD-Cre 2.0 is strongly degraded as compared with the original DD-Cre. On the other hand, DD-Cre 2.0 is quickly stabilized in the presence of TMP in a dose-dependent manner, thereby inducing chemical-dependent DNA recombination. This robust technique further advances applications for conditional genome engineering in living systems.