Fig. 1: Schematic representation of the concept of the Ser/Leu-swapped genetic code with the hybrid-SL tRNA set.

tRNASer, tRNALeu, and tRNATyr are excluded from the E. coli natural tRNA extract. Chimeric tRNALeuGGA, chimeric tRNASerGAG, and tRNATyr synthesized by in vitro transcription are added to the tRNASer, tRNALeu, and tRNATyr-excluded extract to prepare a hybrid-SL tRNA set. Using this hybrid tRNA set, a cell-free translation system with the Ser/Leu-swapped genetic code is constructed, where Ser is assigned to Leu codons and Leu is assigned to Ser codons. A functional protein is synthesized from a Ser/Leu-swapped gene of interest. Abbreviations: IVT in vitro transcribed, E. coli Escherichia coli.