Abstract
We describe here a metabolic labeling protocol for detecting cellular transcriptome-wide mRNA N6 -methyladenosine (m6A) at base resolution. By feeding cells an analog of methionine, potential mRNA m6A forming sites are replaced with the N6 -allyladenosine (a6A). A mild chemical iodination of a6A in RNA results in its opposite base misincorporation during RNA reverse transcription, and thus m6A locations could be precisely identified in the high-throughput sequencing. m6A-label-seq provides a strategy to label and identify cellular epitranscriptomic modification sites. For complete details on the use and execution of this profile, please refer to Shu et al., 2020.
Keywords:
RNA-seq; cell culture; chemistry; molecular biology; molecular/chemical probes; sequence analysis; sequencing.