Chemical probe-based Nanopore direct RNA sequencing method to selectively assess the pseudouridine and inosine modifications
Current methods to identify RNA modifications with short-read sequencing are laborious and direct RNA sequencing gets proclaimed as the viable alternative. Herein, we harness the selective reactivity of the acrylonitrile towards the Inosine (I) and pseudouridine ({Psi}) modifications and developed a chemical probe-based direct RNA sequencing method. We first demonstrated that the chemical probe-induced differential signature profile using nanopore sequencing could facilitate the selective assessment of I and{Psi} in the in vitro synthesized RNA. Furthermore, we verified the I and{Psi} modification with single-nucleotide resolution using RNA derived from mouse brain without the need for a null dataset using knockouts. Our chemical probe-based nanopore sequencing strategy can be extended to profile multiple RNA modifications on a single RNA and may facilitate the diagnosis of disease-associated epitranscriptome markers by generating a comparative dataset in clinical scenarios. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=39 SRC="FIGDIR/small/105338v3_ufig1.gif" ALT="Figure 1"> View larger version (10K): org.highwire.dtl.DTLVardef@137799aorg.highwire.dtl.DTLVardef@110764aorg.highwire.dtl.DTLVardef@1c61ddaorg.highwire.dtl.DTLVardef@1d1db4d_HPS_FORMAT_FIGEXP M_FIG C_FIG