m5C MeRIP sequencing in PANC-1 cells to map the m5C methylome of PANC-1 cells treated with CAF-derived Evs

EV-packaged PIAT mediates EGR1, NTRK1 and SMAD7 mRNA stability in a YBX1-dependent manner. To explore whether YBX1 enhances EGR1, NTRK1 and SMAD7 mRNA stability in an m5C-dependent manner, we conducted m5C-seq in PANC-1 cells to map the m5C methylome of PANC-1 cells treated with CAF-derived EVs. PANC-1 cells were pretreated with CAFs-derived Evs and collected for m5C MeRIP-seq. Briefly, m5C RNA immunoprecipitation was performed with the GenSeqTM m5C RNA IP Kit (GenSeq Inc., China) by following the manufacturer’s instructions. Both the input samples without immunoprecipitation and the m5C IP samples were used for RNA-seq library geneMouseion with NEBNext® Ultra II Directional RNA Library Prep Kit (New England Biolabs, Inc., USA). The library quality was evaluated with BioAnalyzer 2100 system (Agilent Technologies, Inc., USA). Library sequencing was performed on an illumina Novaseq 6000 instrument with 150bp paired-end reads.