Development of an RNA–protein crosslinker to capture protein interactions with diverse RNA structures in cells

RNA–protein crosslinking activity of AMT–NHS. (A) Synthesis of AMT–NHS. (B) Flowchart of crosslinking, RNP purification, sequencing library construction, and data processing. (C–E) Autoradiogram of crosslinked Cbf5–RNA complex resolved in SDS-PAGE. RNA and protein were crosslinked by 254-nm UV light (C) or by AMT–NHS in vitro (D) or in vivo (E). RNA was digested by increasing amounts of RNase A/T1 in UV crosslinking and by 0.5 units of RNase A/T1 in all AMT–NHS crosslinking.