Skip to main content
. 2021 Aug 19;12:5031. doi: 10.1038/s41467-021-25323-x

Fig. 1. Construction and validation of saFRET biosensors.

Fig. 1

a Schematics of mammalian cell biosensor library development, screening, and sequencing in mammalian cells. b Domain structure and activation mechanism of a saFRET biosensor with a fused kinase domain. c–d Representative images (c) and time courses (d) of Fyn-saFRET biosensor with active kinase domain (KA) (n = 98) or kinase-dead domain (KD) (n = 69) before and after PP1 treatment. Error bars, mean ± SD. Scale bars, 10 µm. The color bar indicates ECFP/FRET emission ratio, with hot and cold colors representing the high and low ratios, respectively. e Quantification of the basal FRET ratio of Fyn-saFRET biosensor with KA (n = 98) or KD domain (n = 69). (Unpaired two-tailed Student’s t-test, ****P < 0.0001). Error bars, mean ± SD. f Western blot analysis of the biosensor phosphorylations. Each biological replicate had similar results (n = 3). g Modularized template for the library generation of biosensor variants. The bottom panel illustrates the PCR product of substrate variants using the NNK primer. Source data are provided as a Source Data file.