Infrared adapter ligation allows visualization of RNA–protein complexes with high sensitivity. (A) Schematic of the DNA adapter. The preadenylated DNA adapter contains the UMI, sample barcode, and Illumina adapter sequence. The IR800CW dye is azide-conjugated to the 3′end of the adapter. (B) The infrared adapter can be detected at the TBE-Urea gel starting from 100 attomoles (∼1 pg DNA). (C) SDS–PAGE of the RNA–protein complexes after infrared adapter ligation. Immunoprecipitation of endogenous MOV10 (114 kDa) or IGF2BP3 (64 kDa) proteins was done in 4sU-crosslinked lysates. The control without crosslinking shows a gel background. The crosslinked RNA-adapter fragments add ∼30–50 kDa to the apparent molecular weight of the protein depending on the RNase treatment conditions.
