The nonsense-mediated RNA decay (NMD) pathway is a crucial mechanism of mRNA quality control. Current annotations of NMD substrate RNAs are rarely data-driven, but use general established rules. We present a dataset with 4 cell lines and combinations for SMG5, SMG6 and SMG7 knockdowns or SMG7 knockout. Based on this dataset, we implemented a workflow that combines Nanopore and Illumina sequencing to assemble a transcriptome, which is enriched for NMD target transcripts. Moreover, we use …