Recent Developments in tRNA Sequencing Methods as Applied to Analyses of Mitochondrial tRNAs

Abstract

With the extensive literature that has already accumulated on tRNAs, it may not be evident that this is a relatively new area of research. Discovered only about 22 years ago (Hoagland et al. 1957), following the prediction of the existence of molecules with similar properties by Crick, the first sequence of a tRNA and indeed of any nucleic acid was published only in 1965 (Holley et al. 1965). Since then the sequence of at least 120 different tRNAs from a variety of biological sources have been established (Sprinzl et al. 1978). The two factors that have contributed most to this rapid progress in tRNA sequencing have been the development (1) of column chromatographic and gel electrophoretic methods (Gillam et al. 1967; Cherayil and Bock 1965; Pearson et al. 1971; Ikemura and Dahlberg 1973) suitable for purification of tRNAs and (2) of rapid sequencing methods requiring only very small amounts of tRNAs. Thus, whereas the first sequence analysis of a tRNA, which utilized spectrophotometric procedures for identification of nucleotides (Holley 1968), required several hundred milligrams to a gram of purified tRNA and several years of effort, methods currently available enable one to sequence a tRNA with just a few micrograms (2–10 μg) within a few weeks.

The first few tRNAs to be sequenced were all from yeast (Holley et al. 1965; Zachau et al. 1966; Madison et al. 1966; RajBhandary et al. 1967b; Baev et al. 1967). To a large extent this was a reflection of yeast being a relatively...