A chemically inducible, leakless Cre recombinase by split-protein-based efficient and enhanced degradation (SPEED)

Chemically inducible DNA recombination systems are a very attractive tool for implementing potent genetic applications. However, conventional systems still suffer from leaky properties in the absence of chemicals. Here, we describe a chemically inducible, leakless destabilized Cre recombinase (SPEED-Cre) based on a new approach, split-protein-based efficient and enhanced degradation (SPEED), that consists of a self-assembling split-Cre tagged with a destabilizing domain (DD) mutant from the Escherichia coli dihydrofolate reductase that is stabilized by the antibiotic ligand trimethoprim (TMP). We demonstrate that SPEED-Cre has no significant leak activity of background DNA recombination in the absence of TMP; nevertheless, it enables full induction of TMP-dependent Cre-loxP recombination in human cells and living mice. We also demonstrate the general applicability of the SPEED approach, which can be widely applied to other proteins, by showing high TMP-dependent recombination performances of destabilized Flp, VCre, and Dre recombinases based on the SPEED approach. This robust platform technology will greatly enhance chemogenetic applications for genome engineering in living systems.