m6A-seq data analysis of control and PCIF1 knockdown transcriptome

Purpose: To identify the in vivo RNA targets of PCIF1, we performed meRIP experiments for PCIF1 KD and control cells using an anti-m6A antibody. We envisioned that the cap-specific PCIF1 should selectively alter the m6Am modification at the 5-terminal region of transcripts. Indeed, we observed a reduction of modification peaks at the 5’-end but not the 3’-UTR regions of mRNA upon PCIF1 KD. Methods: Immunoprecipitated RNA fragments were subjected to library construction using NEBNext® UltraTM Directional RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations. Library sequencing was performed on llumina Hiseq X10 with paired-end 2x150 bp read length. Processed reads were mapped to human genome (hg19, UCSC Genome Browser) using tophat2 aligner (v2.0.13) with default parameters.