(A) Full-length mature tRNAs; (B) 5'-tRNA halves; (C) 3'-tRNA halves; and (D) 3'-tRFs. The tRNA name and length are indicated at the top, with an arrow indicating the 5' to 3' orientation, and the tRNA sequence with anticodon positions boxed in red shown below. The IGV coverage plots and alignments below the tRNA sequence were generated from combined datasets for DNase I-treated plasma RNA (n = 12) mapped to mature tRNA reference sequences containing a 3' CCA tail (Chan and Lowe, 2016). Coverage at each position along the gene is shown in the top track, and read alignments are shown below with reads down sampled to a maximum of 100 reads for display when necessary. Gray represents bases that match the reference base. Other colors indicate bases that do not match the reference base (red, green, blue, and brown indicate thymidine, adenosine, cytidine, and guanosine, respectively). Misincorporation at known sites of post-transcriptionally modified bases are highlighted in the alignments: red boxes, m1A58: 1-methyladenosine at position 58; black boxes, m1G37 in His-GTG, m1G9 in iMet-CAT, Cm (2’O-methylation of ribose at C34) in Val-CAC, and −1G in His-GTG. Because tRNA halves and tRFs are much less abundant than mature tRNAs, for visualization in panels B-D, only read pairs with read span < 50 nt and a 3'-RNA end mapping to the anticodon loop are shown for 5’-tRNA halves, and only read pairs with a read span < 50 nt and ending with a 3'-CCA tail are shown for 3'-tRNA halves and 3'-tRFs. Reads corresponding to 3’-tRFs are in orange boxes, and 3’ CCAs are in green boxes. NTA/black boxes are non-templated nucleotides added by TGIRT-III to the 3' end of cDNAs (5' end of the RNA sequence) during TGIRT-seq library preparation.