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Objectives <p>CRISPR-Cas9 nucleases are widely used to introduce targeted DNA double-strand breaks (DSBs) for genome engineering, but the long-term impact of these lesions on local epigenetic information remains poorly characterized. In a companion research article, we used Cas9-assisted targeted nanopore sequencing (CTS) to reveal that CRISPR–Cas9–induced DSBs can disrupt local epigenetic maintenance across multiple genomic contexts and cell systems. Here, we present a structured description of the raw and minimally processed datasets underlying the study. These datasets provide base-resolution measurements of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) at the differentially methylated regions (DMRs) of several imprinted loci, two heterochromatic regions, a cancer-associated promoter epimutation region, and the <i>SNRPN</i> DMR at early/late passages of a clonal line. They enable re-analysis and methodological benchmarking of DSB-associated epigenetic instability.</p> Data description <p>We provide aligned BAM files and per-CpG methylation calls for multiple genomic contexts under both CRISPR-targeted and non-targeting control conditions. Specifically, the collection includes: (i) imprinted loci in human embryonic stem cells (hESCs), including <i>small nuclear ribonucleoprotein polypeptide N</i> (<i>SNRPN</i>), <i>paternally expressed 10</i> (<i>PEG10</i>), and <i>KCNQ1 opposite strand/antisense transcript 1</i> (<i>KCNQ1OT1</i>), (ii) heterochromatic regions in hESCs, including <i>urothelial cancer associated 1</i> (<i>UCA1)</i>, and <i>cysteine rich C-terminal 1 (CRCT1)</i>), (iii) the epimutation locus of <i>MutL homolog 1 (MLH1)</i> in RKO cells, and (iv) the DMR of <i>SNRPN</i> locus in early- and late-passage derivatives of a single hESC clone. For each collection, there is a dataset that includes both the raw aligned Nanopore sequencing reads (BAM) deposited in the NCBI Sequence Read Archive (SRA) and the corresponding processed per-CpG 5mC/5hmC matrices deposited in Zenodo. All higher-level analyses in the research article–such as DMR calling, haplotype-resolved analyses, and structural variant (SV) characterization–are fully reproducible using these deposited data. Additional processed analyses are comprehensively documented in the companion article and are therefore not duplicated here. Together, these datasets offer a rich resource for benchmarking long-read methylation analysis workflows and further investigation of DSB-associated epigenetic instability across diverse genomic contexts.</p>

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Targeted native long-read sequencing of DNA methylation alterations following CRISPR-Cas9-induced double-strand breaks in human cells

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年3月10日

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  • Yingzi Zhang,
  • Mengge Wang,
  • Chongwei Bi,
  • Mo Li

摘要

Objectives

CRISPR-Cas9 nucleases are widely used to introduce targeted DNA double-strand breaks (DSBs) for genome engineering, but the long-term impact of these lesions on local epigenetic information remains poorly characterized. In a companion research article, we used Cas9-assisted targeted nanopore sequencing (CTS) to reveal that CRISPR–Cas9–induced DSBs can disrupt local epigenetic maintenance across multiple genomic contexts and cell systems. Here, we present a structured description of the raw and minimally processed datasets underlying the study. These datasets provide base-resolution measurements of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) at the differentially methylated regions (DMRs) of several imprinted loci, two heterochromatic regions, a cancer-associated promoter epimutation region, and the SNRPN DMR at early/late passages of a clonal line. They enable re-analysis and methodological benchmarking of DSB-associated epigenetic instability.

Data description

We provide aligned BAM files and per-CpG methylation calls for multiple genomic contexts under both CRISPR-targeted and non-targeting control conditions. Specifically, the collection includes: (i) imprinted loci in human embryonic stem cells (hESCs), including small nuclear ribonucleoprotein polypeptide N (SNRPN), paternally expressed 10 (PEG10), and KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1), (ii) heterochromatic regions in hESCs, including urothelial cancer associated 1 (UCA1), and cysteine rich C-terminal 1 (CRCT1)), (iii) the epimutation locus of MutL homolog 1 (MLH1) in RKO cells, and (iv) the DMR of SNRPN locus in early- and late-passage derivatives of a single hESC clone. For each collection, there is a dataset that includes both the raw aligned Nanopore sequencing reads (BAM) deposited in the NCBI Sequence Read Archive (SRA) and the corresponding processed per-CpG 5mC/5hmC matrices deposited in Zenodo. All higher-level analyses in the research article–such as DMR calling, haplotype-resolved analyses, and structural variant (SV) characterization–are fully reproducible using these deposited data. Additional processed analyses are comprehensively documented in the companion article and are therefore not duplicated here. Together, these datasets offer a rich resource for benchmarking long-read methylation analysis workflows and further investigation of DSB-associated epigenetic instability across diverse genomic contexts.