I-tRAP (individual tRNA acylation PCR): A convenient method for selective quantification of tRNA charging.

ABSTRACT: Each transfer RNA (tRNA) is aminoacylated (charged) with a genetic codon-specific amino acid at its 3' end. Charged tRNAs are primarily used for translation, whereas fluctuations in charged tRNA fractions are known to reflect cellular response to stress. Here we report the development of individual tRNA-acylation using PCR (i-tRAP), a convenient PCR-based method that can specifically quantify individual tRNA charging ratio. In this i-tRAP method, demethylases remove base methylations which are problematic for reverse transcription reaction, and β-elimination reaction specifically removes the 3' end of adenine residue in uncharged tRNA. Subsequent TaqMan MGB qRT-PCR can distinguish between cDNA of charged tRNA and uncharged tRNA. By using this method, we revealed that the charging ratio of

SUBMITTER: Tsukamoto Y 

PROVIDER: S-EPMC9808569 | biostudies-literature | 2022 Oct

REPOSITORIES: biostudies-literature