The preparation of liquid and solid media was performed as described previously.17 (link) SD-SCAA and SG-SCAA media used here were prepared in the following combinations: without tryptophan (TRP), leucine (LEU), and uracil (URA) (for DPS experiments with pCTCON2 and pRS315); without TRP and URA (for SPS experiments with pCTCON2); without LEU and URA (for DPS experiments with pRS416 and pRS315); or without URA (for SPS experiments with pRS416). Noncanonical amino acid stocks were prepared at 50 mM L-isomer concentrations. DI water was added to the solid ncAA to 90% of the final volume and 6.0 N NaOH was used as needed to fully dissolve ncAA powders. DI water was added to the final volume and the solution was sterile filtered through a 0.2 μm filter. OmeY was pH adjusted to 7 prior to sterile filtering. Filtered solutions were stored at 4°C for up to two weeks except for photolabile AzF, where a 50 mM stock was made within one week of induction. The strain RJY100 was constructed using standard homologous recombination approaches as described previously.61 (link) Yeast strain BY4741 was purchased from Horizon Discovery (previously Dharmacon). BY4741ΔPPQ1 was a gift from the Fuchs Lab at Tufts University.