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Cited by 1DNA Repair MechanismsCarcinogens and Genotoxicity AssessmentGenomics and Phylogenetic Studies

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Precise mapping of single-stranded DNA breaks by sequence-templated erroneous DNA polymerase end-labelling

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Wenson, Leonie, Heldin, Johan, Martin, Marcel, Erbilgin, Yücel, Salman, Barış, Sundqvist, Anders, Schaal, Wesley, Sandbaumhüter, Friederike A., Jansson, Erik T., Chen, Xingqi, Davidsson, Anton, Stenerlöw, Bo, Espinoza, Jaime A., Lindström, Mikael, Lennartsson, Johan, Spjuth, Ola, Söderberg, Ola·2025·Nature Communications
DOI 10.1038/s41467-025-62512-4PMC PMC12322144

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Abstract

The ability to analyze whether DNA contains lesions is essential in identifying mutagenic substances. Currently, the detection of single-stranded DNA breaks (SSBs) lacks precision. To address this limitation, we develop a method for sequence-templated erroneous end-labelling sequencing (STEEL-seq), which enables the mapping of SSBs. The method requires a highly error-prone DNA polymerase, so we engineer a chimeric DNA polymerase, Sloppymerase, capable of replicating DNA in the absence of one nucleotide. Following the omission of a specific nucleotide (e.g., dATP) from the reaction mixture, Sloppymerase introduces mismatches directly downstream of SSBs at positions where deoxyadenosine should occur. This mismatch pattern, coupled with the retention of sequence information flanking these sites, ensures that the identified hits are bona fide SSBs. STEEL-seq is compatible with a variety of sequencing technologies, as demonstrated using Sanger, Illumina, PacBio, and Nanopore systems. Using STEEL-seq, we determine the SSB/base pair frequency in the human genome to range between 0.7 and 3.8 × 10 −6 with an enrichment in active promoter regions.

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Metadata

Authors
Wenson, L., Heldin, J., Martin, M., Erbilgin, Y., Salman, B., Sundqvist, A., Schaal, W., Sandbaumhüter, F. A., Jansson, E. T., Chen, X., Davidsson, A., Stenerlöw, B., Espinoza, J. A., Lindström, M., Lennartsson, J., Spjuth, O., & Söderberg, O.
Affiliations
  • Uppsala University (SE)
  • Stockholm University (SE)
  • Istanbul University (TR)
  • Science for Life Laboratory (SE)
Corresponding author
Söderberg, Ola
ORCID
Venue
Nature Communications
Publisher
Springer Science and Business Media LLC
Published
2025-08-04
Volume
16
Issue
1
Pages
7130-7130
ISSN
2041-1723 (ISSN-L)
DOI
10.1038/s41467-025-62512-4
PMC
PMC12322144
OpenAlex
W4412947224
Cited by
1
References
5
Citations by year
2026: 1
FWCI
0.30
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Communications (publishedVersion, cc-by)
  • Research Square (acceptedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Diva portal (Dalarna University Library) (submittedVersion, other-oa)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
Cancerfonden (22 2306 Pj)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Carcinogens and Genotoxicity Assessment
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Phylogenetic Studies
Keywords
single-strand breaks, error-prone DNA polymerases, mutagen detection, promoter
MeSH terms
DNA-Directed DNA Polymerase, Humans, Nucleic Acid Conformation, Protein Engineering, Sequence Analysis, DNA, DNA Breaks, Single-Stranded

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