Figure 2.

a) Schematic of the pRG reporter plasmid. Upon light activation, the presence of both gRNAs leads to excision of the DsRed-terminator cassette and NHEJ activates expression of EGFP. b) HEK293T cells transfected with the pRG reporter plasmid, followed by delivery of Cas9:gRNA RNP complexes, were irradiated or kept in the dark. EGFP expression was only observed with non-caged gRNAs or when the caged gRNA was photochemically activated (scale bars = 100 μm). c) EGFP and DsRed fluorescence was quantified by integration of fluorescence intensity in three independently transfected and treated wells for each condition using ImageJ. d) Spatial control of light-activated Cas9:gRNA function through patterned irradiation. HEK293T cells transfected with the pRG reporter and the Cas9:caged gRNA complex were exposed to 365 nm irradiation through a 2 mm-wide slit in a mask (scale bar = 100 μm).