In the mid-1960s, the role of tRNA in protein synthesis was being intensively studied.  At this point, ribosomes had been implicated in protein synthesis, and it had been shown that an mRNA strand was necessary for the formation of these structures.  In a 1964 publication, Warner and Rich showed that ribosomes active in protein synthesis contained tRNA molecules bound at the A and P sites, and discussed the notion that these molecules aided in the peptidyl transferase reaction. However, despite considerable biochemical characterization, the structural basis of tRNA function remained a mystery.  In 1965, Holley et al. purified and sequenced the first tRNA molecule, initially proposing that it adopted a cloverleaf structure, based largely on the ability of certain regions of the molecule to form stem loop structures. The isolation of tRNA proved to be the first major windfall in RNA structural biology.  Following Robert W. Holleys publication, numerous investigators began work on isolation tRNA for crystallographic study, developing improved methods for isolating the molecule as they worked.  By 1968 several groups had produced tRNA crystals, but these proved to be of limited quality and did not yield data at the resolutions necessary to determine structure. In 1971, Kim et al. achieved another breakthrough, producing crystals of yeast tRNA that diffracted to 2–3 Ångström resolutions by using spermine, a naturally occurring polyamine, which bound to and stabilized the tRNA.  Despite having suitable crystals, however, the structure of tRNA was not immediately solved at high resolution; rather it took pioneering work in the use of heavy metal derivatives and a good deal more time to produce a high-quality density map of the entire molecule.  In 1973, Kim et al.' produced a 4 Ångström map of the tRNA molecule in which they could unambiguously trace the entire backbone.  This solution would be followed by many more, as various investigators worked to refine the structure and thereby more thoroughly elucidate the details of base pairing and stacking interactions, and validate the published architecture of the molecule.
The tRNA structure is notable in the field of nucleic acid structure in general, as it represented the first solution of a long-chain nucleic acid structure of any kind—RNA or DNA—preceding Richard E. Dickerson's solution of a B-form dodecamer by nearly a decade. Also, tRNA demonstrated many of the tertiary interactions observed in RNA architecture which would not be categorized and more thoroughly understood for years to come, providing a foundation for all future RNA structural research.