Experimental validation of tRF expression.
A and B. Sequence alignment of tRF-5s with their parental mature tRNAs. The most abundantly cloned tRF-5 isoforms, together with their cloning numbers, are shown. For tRF5-ValGTG, our deep sequencing detected a significant quantity of longer isoforms (designated by “tRF5-ValGTG(l)”) and a short isoform (designated by “tRF5-ValGTG(s)”), both of which were detected in our Northern hybridization (panels shown below). In mature tRNAs, arrowheads on the top indicate cleavage sites based on the isoforms. Anticodons are highlighted by grey. “CCA” in parenthesis indicates a CCA sequence that is post-transcriptionally added to the 3'-end of tRNA. The lengths of each tRNA and tRF-5s are indicated in parentheses. Northern probes (used in next panels) are also aligned and shown. C-F. Northern hybridization of each tRF-5. Photographs of an autoradiogram (designated “Northern” at the bottom) and the ethidium bromide-stained gel (designated “EtBr” at the bottom) are shown. Molecular size markers (in nts) and identities of each band are also indicated. Northern experiments were performed on the mouse tissues used in the deep sequencing (panels C-D) and HUVECs after indicated treatments (panels E-F).