{
  "abstract": "Nanopore direct RNA sequencing (DRS) has transformed transcriptomics by enabling single-molecule, long-read sequencing of native RNA without the need for reverse transcription or amplification. In contrast to short-read RNA-seq and cDNA-based long-read approaches, DRS can simultaneously capture multiple RNA modifications, full-length transcript architecture, alternative splicing patterns, and poly(A) tail features within individual molecules, thereby providing an integrated view of transcriptomic and epitranscriptomic regulation. In this comprehensive review, we outline the biophysical principles underlying nanopore DRS and trace its technological evolution. We compare its performance with short-read RNA sequencing, long-read cDNA sequencing, and conventional RNA-modification mapping strategies, highlighting its advantages in isoform-resolved quantification and multilayer RNA feature integration, while also clarifying contexts in which alternative or combined approaches may be more appropriate for robust biological interpretation. We further summarize optimized experimental workflows, including library construction strategies tailored to diverse RNA biotypes (mRNA, rRNA, tRNA, circRNA, miRNA, and nonpoly(A) transcripts), as well as recommended quality-control procedures and sequencing optimization practices. Emphasizing recent computational advances and translational applications of DRS, we cover state-of-the-art algorithms for RNA modification detection, transcript reconstruction, and isoform quantification. We also propose analytical pipelines for poly(A) tail length inference and integrative frameworks that jointly analyze these regulatory layers. We distinguish direct nanopore signals from computational inferences to define confidence levels and emphasize benchmarking and orthogonal validation of readouts. Practical implementation examples are included to facilitate reproducible analysis. Finally, we highlight emerging applications of integrated DRS, including the resolution of complex transcriptomes, the characterization of coordinated epitranscriptomic regulation, and the identification of disease-associated RNA signatures. We also discuss current technical challenges and future perspectives, particularly in relation to multi-omics integration and the broader deployment of DRS in precision medicine as well as in plant and animal research.",
  "abstract_shown": true,
  "affiliations": [
    "Benagen Institute Wuhan China",
    "Department of Urology The First Affiliated Hospital of Xi'an Jiaotong University Xi'an China",
    "School of Mathematics and Statistics Wuhan University Wuhan China",
    "Key Laboratory of Birth Defects and Related Diseases of Women and Children, Ministry of Education, State Key Laboratory of Biotherapy, Department of Laboratory Medicine West China Second University Hospital Sichuan University Chengdu China",
    "School of Basic Medical Sciences and Forensic Medicine, North Sichuan Medical College, Institute of Basic Medicine Nanchong China",
    "School of Life Sciences and Biotechnology Shanghai Jiao Tong University Shanghai China",
    "Precision Research Center for Refractory Diseases, Shanghai Jiao Tong University Pioneer Research Institute for Molecular and Cell Therapies, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine Shanghai China",
    "College of Life Science and Technology Guangxi University Nanning China",
    "Guangxi Key Laboratory of Organ Donation and Transplantation, Guangxi Key Laboratory of AIDS Prevention and Treatment Institute of Transplant Medicine, The Second Affiliated Hospital of Guangxi Medical University, Guangxi Clinical Research Center for Organ Transplantation Guangxi Medical University Nanning China",
    "Institute of Herbgenomics Chengdu University of Traditional Chinese Medicine Chengdu China",
    "School of Pharmacy Chengdu University of Traditional Chinese Medicine Chengdu China",
    "Faculty of Health and Life Sciences INTI International University Nilai Negeri Sembilan Malaysia",
    "Lee Kong Chian School of Medicine Nanyang Technological University Singapore Singapore",
    "School of Chemistry, Chemical Bioneering, and Biotechnology Nanyang Technological University Singapore Singapore",
    "Sailgene Technology Co., Limited Hong Kong China",
    "Biomedical Sciences College & Shandong Medicinal Biotechnology Centre, Shandong First Medical University & Shandong Academy of Medical Sciences Jinan China",
    "Department of Bioinformatics and Biotechnology Government College University Faisalabad Pakistan",
    "CIRAD, UMR AGAP Institut Guadeloupe Petit Bourg France",
    "UMR AGAP Institut, CIRAD, INRAE, Univ Montpellier, Institut Agro Montpellier France",
    "Multi‐omics Platform for Agricultural Organisms Yazhouwan National Laboratory Sanya China",
    "Institute for Math & AI Wuhan University Wuhan China"
  ],
  "article_type": "review",
  "arxiv": null,
  "author_total": 28,
  "authors": [
    {
      "affs": [
        0,
        1
      ],
      "name": "Tianyuan Zhang",
      "orcid": "0000-0001-8968-563X"
    },
    {
      "affs": [
        2
      ],
      "name": "Jia Li",
      "orcid": "0009-0003-6734-2398"
    },
    {
      "affs": [
        3,
        4
      ],
      "name": "Chao Tang",
      "orcid": "0000-0003-2524-8451"
    },
    {
      "affs": [
        5
      ],
      "name": "You Wu",
      "orcid": "0000-0002-5385-4806"
    },
    {
      "affs": [
        6
      ],
      "name": "H Wu",
      "orcid": "0000-0003-1628-0580"
    },
    {
      "affs": [
        7
      ],
      "name": "Zhu Xi-Tong",
      "orcid": "0009-0001-2477-3089"
    },
    {
      "affs": [
        8
      ],
      "name": "Ziyang Luo",
      "orcid": "0009-0003-0188-2367"
    },
    {
      "affs": [
        1
      ],
      "name": "Hang Qin",
      "orcid": "0000-0002-7435-9652"
    },
    {
      "affs": [
        9,
        10
      ],
      "name": "Lishan Ding",
      "orcid": "0009-0009-9215-9548"
    },
    {
      "affs": [
        6
      ],
      "name": "Yu Zeng"
    },
    {
      "affs": [
        11
      ],
      "name": "Shiou Yih Lee",
      "orcid": "0000-0001-9493-1436"
    },
    {
      "affs": [
        12,
        13
      ],
      "name": "Xiaotao Shen",
      "orcid": "0000-0002-9608-9964"
    },
    {
      "affs": [
        2
      ],
      "name": "Shiwen Gao"
    },
    {
      "affs": [
        14
      ],
      "name": "Zhaoyang Tian",
      "orcid": "0009-0004-4023-7819"
    },
    {
      "affs": [
        7
      ],
      "name": "Qian Tang"
    },
    {
      "affs": [
        15
      ],
      "name": "Mian Li",
      "orcid": "0000-0001-6768-1654"
    },
    {
      "affs": [
        16
      ],
      "name": "Muhammad Tahir ul Qamar",
      "orcid": "0000-0003-4832-4250"
    },
    {
      "affs": [
        1
      ],
      "name": "Yang Dong",
      "orcid": "0000-0001-8378-9008"
    },
    {
      "affs": [
        17,
        18
      ],
      "name": "Komivi Dossa",
      "orcid": "0000-0003-4894-6279"
    },
    {
      "affs": [
        1
      ],
      "name": "Yaxuan Zhang",
      "orcid": "0000-0001-8369-2126"
    },
    {
      "affs": [
        0
      ],
      "name": "Hu Chen",
      "orcid": "0009-0005-5017-9234"
    },
    {
      "affs": [
        8
      ],
      "name": "Sanqi An",
      "orcid": "0000-0002-3177-213X"
    },
    {
      "affs": [
        5
      ],
      "name": "Xiang Yu",
      "orcid": "0000-0002-5730-8802"
    },
    {
      "affs": [
        7,
        3,
        19
      ],
      "name": "Lu Chen",
      "orcid": "0000-0002-1083-9729"
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    {
      "affs": [
        20,
        2
      ],
      "name": "Dingjie Wang",
      "orcid": "0000-0002-2890-5712"
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    {
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        6
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      "name": "Shengli Li",
      "orcid": "0000-0001-5430-303X"
    },
    {
      "affs": [
        7,
        3,
        19
      ],
      "name": "Ling‐Ling Chen",
      "orcid": "0000-0002-3005-526X"
    },
    {
      "affs": [
        1
      ],
      "name": "Yanqiang Li",
      "orcid": "0000-0001-6846-6884"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/8pw1h2cada8tf.bib",
  "cited_by": 17,
  "doi": "10.1002/imt2.70136",
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  "id": "8pw1h2cada8tf",
  "issns": [
    "2770-596X",
    "2770-5986"
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  "landing_url": "https://doi.org/10.1002/imt2.70136",
  "language": "en",
  "licence": "CC-BY",
  "licence_label": "CC BY",
  "licence_source": "journal",
  "licence_url": "https://creativecommons.org/licenses/by/4.0/",
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  "pdf_url": null,
  "pmcid": "PMC13614359",
  "pmid": "42799506",
  "published": "2026-06-01",
  "publisher": "Wiley",
  "ris": "https://research.draconitic.com/paper/8pw1h2cada8tf.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T13:26:46Z",
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      "url": "https://doi.org/10.1002/imt2.70136"
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  "subjects": [
    {
      "field": "biochemistry",
      "origin": "openalex:1312|Genomics and Phylogenetic Studies",
      "subfield": "molecular-biology"
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  ],
  "title": "Nanopore direct RNA sequencing and the epitranscriptome: Advances in mapping native RNA landscapes",
  "title_text": "Nanopore direct RNA sequencing and the epitranscriptome: Advances in mapping native RNA landscapes",
  "type": "peer-reviewed",
  "updated_at": "2026-11-08T13:20:41Z",
  "url": "https://research.draconitic.com/paper/8pw1h2cada8tf",
  "venue": "iMeta",
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