Quantitative profiling of N6-methyladenosine at single-base resolution in stem-differentiating xylem of Populus trichocarpa using Nanopore direct RNA sequencing
Yubang Gao, Xuqing Liu, Bizhi Wu, Huihui Wang, Feihu Xi, Markus V. Kohnen, Anireddy S. N. Reddy, Lianfeng Gu
Abstract
Abstract There are no comprehensive methods to identify N 6 -methyladenosine (m 6 A) at single-base resolution for every single transcript, which is necessary for the estimation of m 6 A abundance. We develop a new pipeline called Nanom6A for the identification and quantification of m 6 A modification at single-base resolution using Nanopore direct RNA sequencing based on an XGBoost model. We validate our method using methylated RNA immunoprecipitation sequencing (MeRIP-Seq) and m 6 A-sensitive RNA-endoribonuclease–facilitated sequencing (m6A-REF-seq), confirming high accuracy. Using this method, we provide a transcriptome-wide quantification of m 6 A modification in stem-differentiating xylem and reveal that different alternative polyadenylation (APA) usage shows a different ratio of m 6 A.