Figure 4.

Efficiency of TISs utilizing non-AUG start codons varies linearly between proteins. The efficiencies of TIS sequences were measured individually by flow cytometry with several genetic reporters and were then compared to the efficiency predicted by FACS-seq. A) GFP reporter (R² = 0.79). B) BFP reporter (R² = 0.83). C) Myc-GFP reporter (the N-terminal of c-Myc fused to GFP, R² = 0.97). Reporter constructs were the same as in Figure 1A with GFP substituted with the other reporter genes. All measured efficiencies were normalized to RFP expression. The TIS sequence CACCAUGG has a TIS efficiency of 100. Error bars represent the standard deviation between experimental replicates (N≥3).