BackgroundBCR::ABL1fusion has significant prognostic value and is screened for chronic myeloid leukemia (CML) disease monitoring as a part of routine molecular testing. To overcome the limitations of the current standard real-time quantitative polymerase chain reaction (RQ-PCR), we designed and validated a next-generation sequencing (NGS)-based assay to quantifyBCR::ABL1andABL1transcript copy numbers.MethodsAfter PCR amplification of the target sequence, deep sequencing was performed using an Illumina Nextseq 550Dx sequencer and in-house–designed bioinformatics pipeline. The Next-generation Quantitative sequencing (NQ-seq) assay was validated for its analytical performance, including precision, linearity, and limit of detection, using serially diluted control materials. A comparison with conventional RQ-PCR was performed with 145 clinical samples from 77 patients.ResultsThe limit of detection of the NQ-seq was the molecular response (MR) 5.6 [BCR::ABL10.00028% international scale (IS)]. The NQ-seq exhibited excellent precision and linear range from MR 2.0 to 5.0. The IS value from the NQ-seq was highly correlated with conventional RQ-PCR.ConclusionsWe conclude that the NQ-seq is an effective tool for monitoringBCR::ABL1transcripts in CML patients with high sensitivity and reliability. Prospective assessment of the unselected large series is required to validate the clinical impact of this NGS-based monitoring strategy.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12935-023-02938-2.