m6A-seq analysis of T-ALL cells following FTO silencing

To identify transcripts directly regulated by FTO through m6A modification in T-ALL, we conducted m6A-seq after silencing FTO in KOPTK1 cells Overall design: After silencing FTO in KOPTK1 cells, total RNA extraction was performed using Trizol. Genomic DNA was eliminated through in-solution DNase I treatment (Thermo Scientific). Subsequently, poly(A) mRNA was purified from the total RNA using the Dynabeads mRNA purification kit (Invitrogen) and randomly fragmented with RNA fragmentation reagents (Ambion). m6A antibody (Abcam) was used to immunoprecipitate m6A RNA.RNA libraries were then constructed using the NEB Next Ultra Directional RNA Library Prep Kit (NEB) and subjected to sequencing as 150 bp paired-end reads on the NovaSeq 6000 platform.