The standard genetic code translates 61 codons into 20 amino acids using fewer than 61 transfer RNAs (tRNAs). This is possible because of the tRNAs ability to wobble at the third base to decode more than one codon. Although the anticodon-codon mapping of tRNA to mRNA is a prerequisite for certain codon usage indices and can contribute to the understanding of the evolution of alternative genetic codes, it is usually not determined experimentally because such assays are prohibitively expensive and elaborate. Instead, the codon reading is approximated from theoretical inferences of nucleotide binding, the wobble rules. Unfortunately, these rules fail to capture all of the nuances of codon reading. This study addresses the codon reading properties of tRNAs and their evolutionary impact on codon usage bias.