Developing a complete set of tRNAs to expand translation control

Previous work has shown that in extraordinary circumstances, the translation of mRNA transcripts with non-canonical start codons can occur in nature through wobble base and near cognate base pairing. Within this thesis, I build upon current bioengineering efforts and create and test a complete set of engineered initiator tRNAs (i-tRNAs) with altered anticodon regions to initiate translation from non-canonical start codons in Escherichia coli. Using bulk fluorescence, bulk luminescence, flow cytometry, targeted mass spectrometry, in silico modelling and fitness assays this thesis highlights the potential utility of using orthogonal i-tRNA mRNA pairs to further the scope of biobased platforms.