Transfer RNAs (tRNAs) are among the most heavily modified RNA species. Posttranscriptional tRNA modifications (ptRMs) play fundamental roles in modulating tRNA structure and function, and are being increasingly linked to human physiology and disease. Detection of ptRMs is often challenging, expensive and laborious. Restriction Fragment Length Polymorphism (RFLP) analyses study the patterns of DNA cleavage after restriction enzyme treatment; and have been used for the qualitative detection of modified bases on messenger RNAs. It is known that some ptRMs induce specific and reproducible base 'mutations' when tRNAs are reverse transcribed. For example, inosine, which derives from the deamination of adenosine, is detected as a guanosine when an inosine-containing tRNA is reverse transcribed, PCR-amplified and sequenced. ptRMdependent base changes on RT-PCR amplicons generated as a consequence of the reverse transcription reaction might create or abolish endonuclease restriction sites. The suitability of RFLP for the detection and/or quantification of ptRMs has not been studied thus far. Here we show that different ptRMs can be detected at specific sites of different tRNA types by RFLP. For the examples studied we show that this approach is able to reliably estimate the modification status of the sample, a feature that can be useful in the study of the regulatory role of tRNA modifications on gene expression.3
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.