Fig. 1: Overview of TECprobe-LM.
From: Sequential structure probing of cotranscriptional RNA folding intermediates
Single-round transcription is initiated on template DNA that contains NPOM-caged-dT and terminal biotin-streptavidin roadblocks. In the initial phase of transcription, RNAP arrests at the NPOM-caged-dT roadblock. The pre-wash sample is removed from the reaction and chemically probed, and the remaining transcription reaction is washed to remove NTPs. The NPOM cage is removed by irradiation with 365 nm UV light (10 mW/cm2 for 3 min) and the pre-chase sample is removed from the reaction and chemically probed. Upon addition of NTPs, RNAP transcribes to the terminal biotin-streptavidin roadblock and the post-chase sample is removed and chemically probed. Figure elements depicting RNA polymerase and nascent RNA were adapted from Szyjka and Strobel, Observation of coordinated RNA folding events by systematic cotranscriptional RNA structure probing20. RNAP, RNA polymerase; SRP, signal recognition particle; SAv, streptavidin; BzCN, benzoyl cyanide.
