tRNA templates were created by in vitro transcription. A list of single-stranded 4 nmol Ultramer DNA oligos (IDT) used as templates are listed in Supplementary Table 1. Each template was designed to harbor a T7 promoter upstream of the tRNA gene sequence. Each template was PCR amplified using a T7 Forward Primer (listed in Supplementary Table 1) and a specific reverse primer complementary to the 3′ end of each tRNA species. PCR amplification was done using Herculase II Fusion DNA Polymerase (Agilent Technologies) following standard procedures. The PCR parameters are as follows: 95 °C for 2 min followed by 35 cycles of 95 °C for 20 s, 48 °C for 20 s and 72 °C for 30 s, ending with 72 °C for 2 min. The PCR products were then resolved on a 2% agarose gel where bands were excised, and DNA was purified using the Qiagen Gel Extraction Kit. In vitro transcription was done using Optizyme T7 RNA Polymerase (ThermoFisher) following standard procedures. Reactions were incubated at 37 °C for 3 h followed by DNase treatment (RQ1 DNase, Promega)
