Rodriguez, J. J. V.; Baxter, K.; Greiss, S.

The ability to precisely control gene expression is fundamental to studying biological processes. Using site-specific recombinases such as Cre or FLP, gene expression can be controlled, albeit with limited spatiotemporal precision, dictated by the expression profile of the promoters used to drive them. Here we develop a photocaged FLP recombinase which can be efficiently and precisely controlled using l ight. To achieve this, we replace critical amino acid residues in the FLP active site with their photocaged counterparts using genetic code expansion technology. We find that photocaged FLP displays no detectable background activity and can be activated by brief illumination in the range of milliseconds to seconds. We further demonstrate that photocaged FLP can be activated using a standard 405 nm microscope-mounted laser, allowing for control of gene expression with single-cell precision.