For the gene sequence isoform (m/z 890, 28-), summaries of identified fragments and their abundances are shown in Figure for the a-EPD, CID, and 213 nm UVPD mass spectra. Sequence coverage above 70% was obtained for all three activation methods, with CID yielding the highest sequence coverage at 83%. Combining the three activation methods yielded near-complete characterization of the tRNA with a total sequence coverage of 97%. These tandem MS results coincided with fragmentation trends from previous studies. CID mainly produced c/y ions and Figure -CID mass spectrum (q 0.14, NCE 25) and sequence coverage map of tRNA-Phe (m/z 890, 28-). Fragments were identified with a 10 ppm m/z tolerance and validated manually. The precursor ion was fully depleted during activation and thus is not labelled. Sequence coverage of 81% was obtained, primarily from c and y fragments. Insets show expansions of isotopic modelling for selected ions from m/z 790 to 795 and 864 to 874. smaller numbers of a/w ions, whereas 213 nm UVPD generated predominantly d and w fragments. For a-EPD, a mix of the ions observed upon CID and UVPD were observed, with d ions being the most often identified and the most abundant. Hydrogen losses were frequently observed in the 213 nm UVPD and a-EPD spectra, most often for d-type ions, as shown in Figure . Hydrogen losses were expected to be prominent for UVPD, as observed in prior studies of proteins and nucleic acids; 10,33 however, single hydrogen losses were often observed for c-type ions across all activation methods. Few complementary pairs of ions were observed for any of the three activation methods, potentially due to the large size of the tRNA and number of possible dissociation pathways, diluting ion fragment ion signal across multiple channels. Furthermore, the potential for "overdissociation" and production of secondary fragments likewise impedes identification of some fragment ions via production of smaller terminal fragments and internal fragments which were not actively considered in this study.