Deciphering transcriptome complexity via long‐read sequencing
Transcriptomics is moving beyond gene-level quantification toward isoform-resolved interrogation of alternative splicing, transcript structural variation, and repeat-derived transcription. Yet short-read sequencing remains intrinsically limited in accurately reconstructing full-length transcripts and resolving complex repetitive regions, including transposable elements. Recent advances in long-read sequencing, exemplified by Pacific Biosciences (PacBio) and Oxford Nanopore Technologies (ONT), offer a transformative opportunity to directly observe complete RNA molecules and thereby overcome these bottlenecks. However, practical adoption is hindered by demanding library construction and the need to process noisy, fast-evolving long-read data, and the field still lacks a unified resource that guides researchers through the entire experimental and analytical workflow. This review fills that gap by providing a concise, end-to-end, and implementation-oriented roadmap for long-read transcriptomics. We distill the key decisions from platform and library strategy selection to core computational processing and downstream interpretation, and we summarize emerging frontiers and best-practice recommendations. By offering a reusable framework and practical checklists, this guide empowers a broader community to exploit long reads for standardized, reproducible, isoform-level discovery at unprecedented resolution.
Authors
- Ana Conesa (ORCID: https://orcid.org/0000-0001-9597-311X)
- Xiufen Zou (ORCID: https://orcid.org/0000-0001-5294-0764)
- Suoqin Jin (ORCID: https://orcid.org/0000-0002-5131-0215)
- Guoqing Tong (ORCID: https://orcid.org/0000-0001-9041-5530)
- Jia Li (ORCID: https://orcid.org/0009-0003-6734-2398)
- Liang Gong (ORCID: https://orcid.org/0000-0003-0563-2579)
- Yilai Han
- Guoliang Chai (ORCID: https://orcid.org/0000-0001-9658-6381)
- Ye Wang (ORCID: https://orcid.org/0000-0002-5423-6196)
- Keying Li (ORCID: https://orcid.org/0000-0003-4934-7489)
- Qian Qin (ORCID: https://orcid.org/0000-0003-3503-3286)
- Yunhao Wang (ORCID: https://orcid.org/0000-0002-6949-290X)
- Dingjie Wang (ORCID: https://orcid.org/0000-0002-2890-5712)
- Wen Hu
- Shiwen Gao
- Tian Wang
- Bo Li
- Chuwen Xu (ORCID: https://orcid.org/0009-0002-3403-2695)
- Tianyuan Zhang
- Yue Yu
- Chenxi Yin (ORCID: https://orcid.org/0009-0006-6521-0967)
- Cheng Chang
Institutions
- Capital Medical University (CN)
- Chinese Academy of Medical Sciences & Peking Union Medical College (CN)
- Chengdu Research Base of Giant Panda Breeding (CN)
- Wuhan University (CN)
- Beijing Geriatric Hospital (CN)
- Stanley Medical Research Institute (US)
- Wuhan Business University (CN)
- First Affiliated Hospital of Xi'an Jiaotong University (CN)
- National Research Council (LK)
- China Academy of Chinese Medical Sciences (CN)
- Shenzhen Institutes of Advanced Technology (CN)
- Beijing Proteome Research Center (CN)
- Institute for Integrative Systems Biology (ES)
- First Affiliated Hospital Zhejiang University (CN)
Publication Details
- Journal
- iMeta
- Published
- 2026-10-04
- DOI
- https://doi.org/10.1002/imt2.70171
- Primary Topic
- Genomics and Phylogenetic Studies
- Type
- article
- Field-Weighted Citation Impact
- 0.00
Funders
- National Natural Science Foundation of China
- National Forestry and Grassland Administration