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. Author manuscript; available in PMC: 2021 Apr 28.
Published in final edited form as: Nature. 2020 Oct 28;587(7835):663–667. doi: 10.1038/s41586-020-2854-z

Fig. 4. Plasmid-encoded cis- and trans-active MTR1.

Fig. 4

a. F30-Broccoli-cis (FBC)-MTR1 (modification site indicated as red dot) is transcribed in E.coli. Total RNA isolated 1h after IPTG induction, analysed on PAGE next to an in vitro transcribed reference and a size marker, and stained by DFHBI (10% denaturing PAGE, 20 μM DFHBI) and Sybr gold. Both the in vitro and in vivo transcripts contain an active MTR1 ribozyme, as revealed by the primer extension stops after incubation with m6G or BG (here with in vitro transcript and primer P10, data with in vivo transcript shown in Extended Data Fig. 7). b. F30-Broccoli-trans (FBT)- MTR1 with binding arms specific for hybridization to E.coli tRNAAsp. The activity was reduced with m6G but retained with BG. Formation of 1-benzyladenosine was confirmed by LC-MS and specifically detected only in tRNAAsp (Extended Data Fig. 8).