InMycoplasma capricolum, codon family boxes except for arginine and threonine (CUN leucine, GUN valine, UCN serine, CCN proline, GCN alanine, and GGN glycine) have only a single tRNA species with the anticodon sequence UNN (N is U,C,A or G), the first nucleoside U being modified. Incorporation of the [ 3 H] amino acid into the peptide fraction was examined with theM. capricolumcell-free translation system. Synthetic mRNA containing each of the respective amino acid codons in the coding frame was subjected to translation. The tRNA U N N translated all the family box codons with similar, if not equal, efficiencies. InM. capricolum, there are two species of threonine tRNA, tRNA U G U T h r and tRNA A G U T h r , the first nucleoside U or A being unmodified. The tRNA U G U T h r species translates codons ACA, ACG and ACU efficiently, and ACC only poorly. In contrast, the tRNA A G U T h r species translates codons ACU, ACC and ACG efficiently and ACA poorly.