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. Author manuscript; available in PMC: 2016 Mar 2.
Published in final edited form as: Methods Enzymol. 2015 Jul 17;560:29–71. doi: 10.1016/bs.mie.2015.03.004

Figure 6.

Figure 6

Quantitative mapping of modified ribonucleosides in a specific tRNA. To assess stress-induced changes in m5C levels in tRNALeu(CAA), two RNase-generated oligonucleotides possessing m5C-modified sites in this tRNA were identified using the Mongo Oligo Mass Calculator. RNase T1 yields a 4-mer oligonucleotide harboring either C or m5C at position 48 (CAAG), while RNase U2 produces a 5-mer oligonucleotide with C or m5C at the wobble position 34 of (UUCAA). The UUCAA oligonucleotide was sequenced and quantified by LC-MS/MS analysis with affinity-purified tRNA and the CAAG oligonucleotide was analyzed by LC-MS/MS analysis using total tRNA.