Long-read DNA and RNA Sequencing Special Collection

Genome Research presents this collection comprising Part 1 and Part 2 of the Special Issue on Long-read DNA and RNA Sequencing Applications in Biology and Medicine, Guest Edited by Drs. Ana Conesa, Alexander Hoischen, and Fritz Sedlazeck, and related research and review articles highlighting novel applications and developments in long-read sequencing. Articles in these issues focus on original research offering novel biological and clinical insights gained using long-read DNA and RNA sequencing technologies and other long molecule approaches. This collection also offers significant advances in long-read sequencing analysis, including novel methods for genome assembly and annotation, characterization of complex structural variation, prioritization of disease variants, DNA and RNA modification, and mRNA and protein isoform diversity. Advances in vaccine and gene therapy, tumor cell-free DNA profiling, bacterial surveillance, and single-cell and spatial long-read sequencing are also featured. Reviews and Perspectives cover the impact and advantages of using long-read sequencing, the remaining challenges, and include advances and insights in technology, rare disease, cancer, population genomics, transcriptomics, and epigenomics.

Illustration by Alex Cagan, University of Cambridge.

Meet the Guest Editors

Ana Conesa is Research Professor at the Institute for Integrative Systems Biology of the Spanish National Research Council (CSIC). She is member Spanish Royal Academy of Engineer, Fellow of the International Society for Computational Biology, honorary member of the Spanish Society of Bioinformatics and Computational Biology. She coordinates the CSIC Connection for Computational Biology and Bioinformatics. Dr. Conesa’s lab is renowned for developing computational tools that facilitate the analysis and integration of transcriptomics and multi-omics data, with a strong focus on uncovering the functional aspects of gene expression on a genome-wide scale. Among her group’s widely-used tools are Blast2GO, PaintOmics, maSigPro, NOISeq, and Qualimap. Her team has been at the forefront of creating tools for long-read transcriptomics analysis and co-leads the international consortium LRGASP, dedicated to benchmarking long-read methods for transcript identification and quantification. Currently, she leads the LongTREC EU consortium, aimed at advancing next-generation bioinformatics tools to support the transition to long-read RNA sequencing. She is also the co-founder of Biobam Bioinformatics, a company that offers user-friendly software solutions for biologists.

Alexander Hoischen’s heads the research group ‘Genomic Technologies for Immune-Mediated and Infectious Diseases’ at Radboud University Medical Center, which builds expertise in the identification of rare disease genes using the latest genomics tools. His group was the first to identify a disease-causing dominant de novo mutation for a Mendelian disorder by exome sequencing, followed by several disease gene discoveries. Recent efforts particularly include applications of long-read sequencing and optical genome mapping to undiagnosed rare disease cases. A new focus are the genetic basis of immune diseases, with the most recent identification of a novel immunodeficiency that predisposes men to severe COVID-19, and a missing SCID gene. Dr. Hoischen co-leads a work package in the EU-funded H2020 project SOLVE-RD (www.solve-rd.eu), which will be followed-up by his work package–lead in the newly funded ERDERA project (https://erdera.org), both with strong focus of long-read sequencing efforts in the rare disease space. Most recently he has received a VICI grant “SOLVE-IEI - Solving Enigmas of Undiagnosed Inborn Errors of Immunity”. Dr. Hoischen and his local, national, and international network of colleagues and collaborators will continue to pioneer novel and disruptive technologies that allow new scientific insights and rapid translation into clinical and diagnostic practice.

Fritz Sedlazeck is an Associate Professor at Baylor College of Medicine and an adjunct Associate Professor at Rice University. He is leading a research group since 2017 at the Human Genome Sequencing Centre at Baylor College of Medicine. His research focuses on developing computational methods to detect and analyze genomic variations with a focus on Structural Variations. Structural Variations are genomic events that manipulate multiple positions in a genome, which impact evolution, genomic disorders, regulation as well as play an important role in explaining multiple phenotypes. Dr. Sedlazeck groups focuses on the mechanisms of the formation of SV across multiple species and to improve our understanding how these complex alleles evolve and impact phenotypes. Dr Sedlazeck’s work has significantly advanced the field of structural variant detection, enabling deeper insights into mosaicism, somatic mutations, and disease-associated genetic variation. He has authored numerous impactful publications and contributed to widely-used bioinformatics tools, positioning him as a leading figure in the application of bioinformatics to genomics and precision medicine.

Long read DNA and RNA Sequencing Special Issue: Part 2

April 2025; 35 (4)

This Part 2 of the Special Issue on Long-read DNA and RNA Sequencing Applications in Biology and Medicine offers significant advances in long-read sequencing analysis, including novel methods for genome assembly and annotation, prioritization of structural variants in rare disease, characterization of complex repeats, RNA modification, mRNA and protein isoform diversity, tumor cell-free DNA profiling, and bacterial surveillance. Reviews and Perspectives include a Hitchhiker’s Guide to long-read genomic analysis, and insights into the additional diagnostic yield of long-read sequencing in undiagnosed rare diseases, the complexity of cancer, the challenges in genome annotation, population genomics, transcriptomics, and epigenomics.

Editorial

  • Long reads decipher genomes and transcriptomes and offer novel insights into biology and diseases

    • Ana Conesa,
    • Alexander Hoischen,
    • and Fritz J. Sedlazeck
    Genome Res.April 2025 35: xi-xiv; doi:10.1101/gr.280665.125

Mini-Reviews

  • A Hitchhiker's Guide to long-read genomic analysis

    • Medhat Mahmoud,
    • Daniel P. Agustinho,
    • and Fritz J. Sedlazeck
    Genome Res. April 2025 35: 545-558; doi:10.1101/gr.279975.124

  • The additional diagnostic yield of long-read sequencing in undiagnosed rare diseases

    • Giulia F. Del Gobbo,
    • and Kym M. Boycott
    Genome Res. April 2025 35: 559-571; doi:10.1101/gr.279970.124

  • Evolution of genome-wide methylation profiling technologies

    • Carolina Montano,
    • and Winston Timp
    Genome Res. April 2025 35: 572-582; doi:10.1101/gr.278407.123

Perspectives

  • Notable challenges posed by long-read sequencing for the study of transcriptional diversity and genome annotation

    • Carolina Monzó,
    • Adam Frankish,
    • and Ana Conesa
    Genome Res. April 2025 35: 583-592; doi:10.1101/gr.279865.124

  • The impact of long-read sequencing on human population-scale genomics

    • Tobias Rausch,
    • Tobias Marschall,
    • and Jan O. Korbel
    Genome Res. April 2025 35: 593-598; doi:10.1101/gr.280120.124

  • Unraveling the hidden complexity of cancer through long-read sequencing

    • Qiuhui Li,
    • Ayse G. Keskus,
    • Justin Wagner,
    • Michal B. Izydorczyk,
    • Winston Timp,
    • Fritz J. Sedlazeck,
    • Alison P. Klein,
    • Justin M. Zook,
    • Mikhail Kolmogorov,
    • and Michael C. Schatz
    Genome Res. April 2025 35: 599-620; doi:10.1101/gr.280041.124

Research

  • Closing the gaps, and improving somatic structural variant analysis and benchmarking using CHM13-T2T

    • Luis F. Paulin,
    • Jeremy Fan,
    • Kieran O'Neill,
    • Erin Pleasance,
    • Vanessa L. Porter,
    • Steven J.M. Jones,
    • and Fritz J. Sedlazeck
    Genome Res. April 2025 35: 621-631; doi:10.1101/gr.279352.124

  • Assessing DNA methylation detection for primary human tissue using Nanopore sequencing

    • Rylee Genner,
    • Stuart Akeson,
    • Melissa Meredith,
    • Pilar Alvarez Jerez,
    • Laksh Malik,
    • Breeana Baker,
    • Abigail Miano-Burkhardt,
    • CARD-long-read Team,
    • Benedict Paten,
    • Kimberley J. Billingsley,
    • Cornelis Blauwendraat,
    • and Miten Jain
    Genome Res. April 2025 35: 632-643; doi:10.1101/gr.279159.124

  • Expanded methylome and quantitative trait loci detection by long-read profiling of personal DNA

    • Cristian Groza,
    • Bing Ge,
    • Warren A. Cheung,
    • Tomi Pastinen,
    • and Guillaume Bourque
    Genome Res. April 2025 35: 644-652; doi:10.1101/gr.279240.124

  • Rearrangements of viral and human genomes at human papillomavirus integration events and their allele-specific impacts on cancer genome regulation

    • Vanessa L. Porter,
    • Michelle Ng,
    • Kieran O'Neill,
    • Signe MacLennan,
    • Richard D. Corbett,
    • Luka Culibrk,
    • Zeid Hamadeh,
    • Marissa Iden,
    • Rachel Schmidt,
    • Shirng-Wern Tsaih,
    • Carolyn Nakisige,
    • Martin Origa,
    • Jackson Orem,
    • Glenn Chang,
    • Jeremy Fan,
    • Ka Ming Nip,
    • Vahid Akbari,
    • Simon K. Chan,
    • James Hopkins,
    • Richard A. Moore,
    • Eric Chuah,
    • Karen L. Mungall,
    • Andrew J. Mungall,
    • Inanc Birol,
    • Steven J.M. Jones,
    • Janet S. Rader,
    • and Marco A. Marra
    Genome Res. April 2025 35: 653-670; doi:10.1101/gr.279041.124

  • Multisite long-read sequencing reveals the early contributions of somatic structural variations to HBV-related hepatocellular carcinoma tumorigenesis

    • Tianfu Zeng,
    • Haotian Liao,
    • Lin Xia,
    • Siyao You,
    • Yanqun Huang,
    • Jiaxun Zhang,
    • Yahui Liu,
    • Xuyan Liu,
    • and Dan Xie
    Genome Res. April 2025 35: 671-685; doi:10.1101/gr.279617.124

  • Long-read single-cell RNA sequencing enables the study of cancer subclone-specific genotypes and phenotypes in chronic lymphocytic leukemia

    • Gage S. Black,
    • Xiaomeng Huang,
    • Yi Qiao,
    • Philip Moos,
    • Deepa Sampath,
    • Deborah M. Stephens,
    • Jennifer A. Woyach,
    • and Gabor T. Marth
    Genome Res. April 2025 35: 686-697; doi:10.1101/gr.279049.124

  • Heterogeneous and novel transcript expression in single cells of patient-derived clear cell renal cell carcinoma organoids

    • Tülay Karakulak,
    • Natalia Zajac,
    • Hella Anna Bolck,
    • Anna Bratus-Neuenschwander,
    • Qin Zhang,
    • Weihong Qi,
    • Debleena Basu,
    • Tamara Carrasco Oltra,
    • Hubert Rehrauer,
    • Christian von Mering,
    • Holger Moch,
    • and Abdullah Kahraman
    Genome Res. April 2025 35: 698-711; doi:10.1101/gr.279345.124

  • Genetic regulation of nascent RNA maturation revealed by direct RNA nanopore sequencing

    • Karine Choquet,
    • Louis-Philippe Chaumont,
    • Simon Bache,
    • Autum R. Baxter-Koenigs,
    • and L. Stirling Churchman
    Genome Res. April 2025 35: 712-724; doi:10.1101/gr.279203.124

  • Deciphering the largest disease-associated transcript isoforms in the human neural retina with advanced long-read sequencing approaches

    • Merel Stemerdink,
    • Tabea Riepe,
    • Nick Zomer,
    • Renee Salz,
    • Michael Kwint,
    • Jaap Oostrik,
    • Raoul Timmermans,
    • Barbara Ferrari,
    • Stefano Ferrari,
    • Alfredo Dueñas Rey,
    • Emma Delanote,
    • Suzanne E. de Bruijn,
    • Hannie Kremer,
    • Susanne Roosing,
    • Frauke Coppieters,
    • Alexander Hoischen,
    • Frans P.M. Cremers,
    • Peter A.C. ‘t Hoen,
    • Erwin van Wijk,
    • and Erik de Vrieze
    Genome Res. April 2025 35: 725-739; doi:10.1101/gr.280060.124

  • Integrating short-read and long-read single-cell RNA sequencing for comprehensive transcriptome profiling in mouse retina

    • Meng Wang,
    • Yumei Li,
    • Jun Wang,
    • Soo Hwan Oh,
    • Yexuan Cao,
    • and Rui Chen
    Genome Res. April 2025 35: 740-754; doi:10.1101/gr.279167.124

  • Unraveling undiagnosed rare disease cases by HiFi long-read genome sequencing

    • Wouter Steyaert,
    • Lydia Sagath,
    • German Demidov,
    • Vicente A. Yépez,
    • Anna Esteve-Codina,
    • Julien Gagneur,
    • Kornelia Ellwanger,
    • Ronny Derks,
    • Marjan Weiss,
    • Amber den Ouden,
    • Simone van den Heuvel,
    • Hilde Swinkels,
    • Nick Zomer,
    • Marloes Steehouwer,
    • Luke O'Gorman,
    • Galuh Astuti,
    • Kornelia Neveling,
    • Rebecca Schüle,
    • Jishu Xu,
    • Matthis Synofzik,
    • Danique Beijer,
    • Holger Hengel,
    • Ludger Schöls,
    • Kristl G. Claeys,
    • Jonathan Baets,
    • Liedewei Van de Vondel,
    • Alessandra Ferlini,
    • Rita Selvatici,
    • Heba Morsy,
    • Marwa Saeed Abd Elmaksoud,
    • Volker Straub,
    • Juliane Müller,
    • Veronica Pini,
    • Luke Perry,
    • Anna Sarkozy,
    • Irina Zaharieva,
    • Francesco Muntoni,
    • Enrico Bugiardini,
    • Kiran Polavarapu,
    • Rita Horvath,
    • Evan Reid,
    • Hanns Lochmüller,
    • Marco Spinazzi,
    • Marco Savarese,
    • Solve-RD DITF-ITHACA, Solve-RD DITF-Euro-NMD, Solve-RD DITF-RND, Solve-RD DITF-EpiCARE,
    • Leslie Matalonga,
    • Steven Laurie,
    • Han G. Brunner,
    • Holm Graessner,
    • Sergi Beltran,
    • Stephan Ossowski,
    • Lisenka E.L.M. Vissers,
    • Christian Gilissen,
    • Alexander Hoischen,
    • and on behalf of the Solve-RD consortium
    Genome Res. April 2025 35: 755-768; doi:10.1101/gr.279414.124

  • A prospective trial comparing programmable targeted long-read sequencing and short-read genome sequencing for genetic diagnosis of cerebellar ataxia

    • Haloom Rafehi,
    • Liam G. Fearnley,
    • Justin Read,
    • Penny Snell,
    • Kayli C. Davies,
    • Liam Scott,
    • Greta Gillies,
    • Genevieve C. Thompson,
    • Tess A. Field,
    • Aleena Eldo,
    • Simon Bodek,
    • Ernest Butler,
    • Luke Chen,
    • John Drago,
    • Himanshu Goel,
    • Anna Hackett,
    • G. Michael Halmagyi,
    • Andrew Hannaford,
    • Katya Kotschet,
    • Kishore R. Kumar,
    • Smitha Kumble,
    • Matthew Lee-Archer,
    • Abhishek Malhotra,
    • Mark Paine,
    • Michael Poon,
    • Kate Pope,
    • Katrina Reardon,
    • Steven Ring,
    • Anne Ronan,
    • Matthew Silsby,
    • Renee Smyth,
    • Chloe Stutterd,
    • Mathew Wallis,
    • John Waterston,
    • Thomas Wellings,
    • Kirsty West,
    • Christine Wools,
    • Kathy H.C. Wu,
    • David J. Szmulewicz,
    • Martin B. Delatycki,
    • Melanie Bahlo,
    • and Paul J. Lockhart
    Genome Res. April 2025 35: 769-785; doi:10.1101/gr.279634.124

  • Multiple paralogs and recombination mechanisms contribute to the high incidence of 22q11.2 deletion syndrome

    • Lisanne Vervoort,
    • Nicolas Dierckxsens,
    • Marta Sousa Santos,
    • Senne Meynants,
    • Erika Souche,
    • Ruben Cools,
    • Tracy Heung,
    • Koen Devriendt,
    • Hilde Peeters,
    • Donna M. McDonald-McGinn,
    • Ann Swillen,
    • Jeroen Breckpot,
    • Beverly S. Emanuel,
    • Hilde Van Esch,
    • Anne S. Bassett,
    • and Joris R. Vermeesch
    Genome Res. April 2025 35: 786-797; doi:10.1101/gr.279331.124

  • Optical genome mapping identifies rare structural variants in neural tube defects

    • Nikhil S. Sahajpal,
    • Jane Dean,
    • Benjamin Hilton,
    • Timothy Fee,
    • Cindy Skinner,
    • Alex Hastie,
    • Barbara R. DuPont,
    • Alka Chaubey,
    • Michael J. Friez,
    • and Roger E. Stevenson
    Genome Res. April 2025 35: 798-809; doi:10.1101/gr.279318.124

  • Optical genome mapping enables accurate testing of large repeat expansions

    • Bart van der Sanden,
    • Kornelia Neveling,
    • Syukri Shukor,
    • Michael D. Gallagher,
    • Joyce Lee,
    • Stephanie L. Burke,
    • Maartje Pennings,
    • Ronald van Beek,
    • Michiel Oorsprong,
    • Ellen Kater-Baats,
    • Eveline Kamping,
    • Alide A. Tieleman,
    • Nicol C. Voermans,
    • Ingrid E. Scheffer,
    • Jozef Gecz,
    • Mark A. Corbett,
    • Lisenka E.L.M. Vissers,
    • Andy Wing Chun Pang,
    • Alex Hastie,
    • Erik-Jan Kamsteeg,
    • and Alexander Hoischen
    Genome Res. April 2025 35: 810-823; doi:10.1101/gr.279491.124

  • Candida albicans isolates contain frequent heterozygous structural variants and transposable elements within genes and centromeres

    • Ursula Oggenfuss,
    • Robert T. Todd,
    • Natthapon Soisangwan,
    • Bailey Kemp,
    • Alison Guyer,
    • Annette Beach,
    • and Anna Selmecki
    Genome Res. April 2025 35: 824-838; doi:10.1101/gr.279301.124

Methods

  • Geometric deep learning framework for de novo genome assembly

    • Lovro Vrček,
    • Xavier Bresson,
    • Thomas Laurent,
    • Martin Schmitz,
    • Kenji Kawaguchi,
    • and Mile Šikić
    Genome Res. April 2025 35: 839-849; doi:10.1101/gr.279307.124

  • Multisample motif discovery and visualization for tandem repeats

    • Yaran Zhang,
    • Marc Hulsman,
    • Alex Salazar,
    • Niccolò Tesi,
    • Lydian Knoop,
    • Sven van der Lee,
    • Sanduni Wijesekera,
    • Jana Krizova,
    • Erik-Jan Kamsteeg,
    • and Henne Holstege
    Genome Res. April 2025 35: 850-862; doi:10.1101/gr.279278.124

  • RAmbler resolves complex repeats in human Chromosomes 8, 19, and X

    • Sakshar Chakravarty,
    • Glennis Logsdon,
    • and Stefano Lonardi
    Genome Res. April 2025 35: 863-876; doi:10.1101/gr.279308.124

  • Enhancing nanopore adaptive sampling for PromethION using readfish at scale

    • Rory Munro,
    • Alexander Payne,
    • Nadine Holmes,
    • Chris Moore,
    • Inswasti Cahyani,
    • and Matthew Loose
    Genome Res. April 2025 35: 877-885; doi:10.1101/gr.279329.124

  • Nanopore-based consensus sequencing enables accurate multimodal tumor cell-free DNA profiling

    • Li-Ting Chen,
    • Myrthe Jager,
    • Dàmi Rebergen,
    • Geertruid J. Brink,
    • Tom van den Ende,
    • Willem Vanderlinden,
    • Pauline Kolbeck,
    • Marc Pagès-Gallego,
    • Ymke van der Pol,
    • Nicolle Besselink,
    • Norbert Moldovan,
    • Nizar Hami,
    • Wigard P. Kloosterman,
    • Hanneke van Laarhoven,
    • Florent Mouliere,
    • Ronald Zweemer,
    • Jan Lipfert,
    • Sarah Derks,
    • Alessio Marcozzi,
    • and Jeroen de Ridder
    Genome Res. April 2025 35: 886-899; doi:10.1101/gr.279144.124

  • De novo detection of somatic variants in high-quality long-read single-cell RNA sequencing data

    • Arthur Dondi,
    • Nico Borgsmüller,
    • Pedro F. Ferreira,
    • Brian J. Haas,
    • Francis Jacob,
    • Viola Heinzelmann-Schwarz,
    • Tumor Profiler Consortium,
    • and Niko Beerenwinkel
    Genome Res. April 2025 35: 900-913; doi:10.1101/gr.279281.124

  • Integration of transcriptomics and long-read genomics prioritizes structural variants in rare disease

    • Tanner D. Jensen,
    • Bohan Ni,
    • Chloe M. Reuter,
    • John E. Gorzynski,
    • Sarah Fazal,
    • Devon Bonner,
    • Rachel A. Ungar,
    • Pagé C. Goddard,
    • Archana Raja,
    • Euan A. Ashley,
    • Jonathan A. Bernstein,
    • Stephan Zuchner,
    • Undiagnosed Diseases Network,
    • Michael D. Greicius,
    • Stephen B. Montgomery,
    • Michael C. Schatz,
    • Matthew T. Wheeler,
    • and Alexis Battle
    Genome Res. April 2025 35: 914-928; doi:10.1101/gr.279323.124

  • De novo antibody identification in human blood from full-length single B cell transcriptomics and matching haplotype-resolved germline assemblies

    • John Beaulaurier,
    • Lynn Ly,
    • J. Andrew Duty,
    • Carly Tyer,
    • Christian Stevens,
    • Chuan-Tien Hung,
    • Akash Sookdeo,
    • Alex W. Drong,
    • Shreyas Kowdle,
    • Axel Guzman-Solis,
    • Domenico Tortorella,
    • Daniel J. Turner,
    • Sissel Juul,
    • Scott Hickey,
    • and Benhur Lee
    Genome Res. April 2025 35: 929-941; doi:10.1101/gr.279392.124

  • Single-cell Rapid Capture Hybridization sequencing reliably detects isoform usage and coding mutations in targeted genes

    • Hongke Peng,
    • Jafar S. Jabbari,
    • Luyi Tian,
    • Changqing Wang,
    • Yupei You,
    • Chong Chyn Chua,
    • Natasha S. Anstee,
    • Noorul Amin,
    • Andrew H. Wei,
    • Nadia M. Davidson,
    • Andrew W. Roberts,
    • David C.S. Huang,
    • Matthew E. Ritchie,
    • and Rachel Thijssen
    Genome Res. April 2025 35: 942-955; doi:10.1101/gr.279322.124

  • Rapid and accurate demultiplexing of direct RNA nanopore sequencing data with SeqTagger

    • Leszek P. Pryszcz,
    • Gregor Diensthuber,
    • Laia Llovera,
    • Rebeca Medina,
    • Anna Delgado-Tejedor,
    • Luca Cozzuto,
    • Julia Ponomarenko,
    • and Eva Maria Novoa
    Genome Res. April 2025 35: 956-966; doi:10.1101/gr.279290.124

  • Accurate fusion transcript identification from long- and short-read isoform sequencing at bulk or single-cell resolution

    • Qian Qin,
    • Victoria Popic,
    • Kirsty Wienand,
    • Houlin Yu,
    • Emily White,
    • Akanksha Khorgade,
    • Asa Shin,
    • Christophe Georgescu,
    • Catarina D. Campbell,
    • Arthur Dondi,
    • Niko Beerenwinkel,
    • Francisca Vazquez,
    • Aziz M. Al'Khafaji,
    • and Brian J. Haas
    Genome Res. April 2025 35: 967-986; doi:10.1101/gr.279200.124

  • Quality assessment of long read data in multisample lrRNA-seq experiments using SQANTI-reads

    • Netanya Keil,
    • Carolina Monzó,
    • Lauren McIntyre,
    • and Ana Conesa
    Genome Res. April 2025 35: 987-998; doi:10.1101/gr.280021.124

  • Long-read RNA sequencing reveals allele-specific N6-methyladenosine modifications

    • Dayea Park,
    • and Can Cenik
    Genome Res. April 2025 35: 999-1011; doi:10.1101/gr.279270.124

  • Biosurfer for systematic tracking of regulatory mechanisms leading to protein isoform diversity

    • Mayank Murali,
    • Jamie Saquing,
    • Senbao Lu,
    • Ziyang Gao,
    • Emily F. Watts,
    • Ben Jordan,
    • Zachary Peters Wakefield,
    • Ana Fiszbein,
    • David R. Cooper,
    • Peter J. Castaldi,
    • Dmitry Korkin,
    • and Gloria M. Sheynkman
    Genome Res. April 2025 35: 1012-1024; doi:10.1101/gr.279317.124

  • Optimizing nanopore adaptive sampling for pneumococcal serotype surveillance in complex samples using the graph-based GNASTy algorithm

    • Samuel T. Horsfield,
    • Basil C.T. Fok,
    • Yuhan Fu,
    • Paul Turner,
    • John A. Lees,
    • and Nicholas J. Croucher
    Genome Res. April 2025 35: 1025-1040; doi:10.1101/gr.279435.124

  • Taurine pangenome uncovers a segmental duplication upstream of KIT associated with depigmentation in white-headed cattle

    • Sotiria Milia,
    • Alexander S. Leonard,
    • Xena Marie Mapel,
    • Sandra Milena Bernal Ulloa,
    • Cord Drögemüller,
    • and Hubert Pausch
    Genome Res. April 2025 35: 1041-1052; doi:10.1101/gr.279064.124

  • Evaluation of strategies for evidence-driven genome annotation using long-read RNA-seq

    • Alejandro Paniagua,
    • Cristina Agustín-García,
    • Francisco J. Pardo-Palacios,
    • Thomas Brown,
    • Maite De Maria,
    • Nancy D. Denslow,
    • Camila J. Mazzoni,
    • and Ana Conesa
    Genome Res. April 2025 35: 1053-1064; doi:10.1101/gr.279864.124
 

Long read DNA and RNA Sequencing Special Issue: Part 1

November 2024; 34 (11)

This Part 1 of the Special Issue on Long-read DNA and RNA Sequencing Applications in Biology and Medicine offers significant advances in long-read sequencing analysis, including novel methods for genome assembly and annotation, characterization of complex structural variation, quantifying DNA and RNA modification, full-length mRNA isoform resolution, vaccine and gene therapy vector quality control, and bacterial outbreak tracing. Reviews of targeted sequencing strategies that leverage existing long-read technologies, the specific challenges involved in identifying and quantifying mRNA terminal ends with these technologies, and advances in single-cell and spatial long-read sequencing are also featured.

Editorial

  • Revolutionizing genomics and medicine—one long molecule at a time

    • Ana Conesa,
    • Alexander Hoischen,
    • and Fritz J. Sedlazeck
    Genome Res. November 2024 34: ix-xi; doi:10.1101/gr.280179.124

Reviews

  • Leveraging the power of long reads for targeted sequencing

    • Shruti V. Iyer,
    • Sara Goodwin,
    • and William Richard McCombie
    Genome Res. November 2024 34: 1701-1718; doi:10.1101/gr.279168.124

  • Understanding isoform expression by pairing long-read sequencing with single-cell and spatial transcriptomics

    • Natan Belchikov,
    • Justine Hsu,
    • Xiang Jennie Li,
    • Julien Jarroux,
    • Wen Hu,
    • Anoushka Joglekar,
    • and Hagen U. Tilgner
    Genome Res. November 2024 34: 1735-1746; doi:10.1101/gr.279640.124

  • Challenges in identifying mRNA transcript starts and ends from long-read sequencing data

    • Ezequiel Calvo-Roitberg,
    • Rachel F. Daniels,
    • and Athma A. Pai
    Genome Res. November 2024 34: 1719-1734; doi:10.1101/gr.279559.124

Research

  • Long-read genome sequencing and variant reanalysis increase diagnostic yield in neurodevelopmental disorders

    • Susan M. Hiatt,
    • James M.J. Lawlor,
    • Lori H. Handley,
    • Donald R. Latner,
    • Zachary T. Bonnstetter,
    • Candice R. Finnila,
    • Michelle L. Thompson,
    • Lori Beth Boston,
    • Melissa Williams,
    • Ivan Rodriguez Nunez,
    • Jerry Jenkins,
    • Whitley V. Kelley,
    • E. Martina Bebin,
    • Michael A. Lopez,
    • Anna C.E. Hurst,
    • Bruce R. Korf,
    • Jeremy Schmutz,
    • Jane Grimwood,
    • and Gregory M. Cooper
    Genome Res. November 2024 34: 1747-1762; doi:10.1101/gr.279227.124

  • Resolving complex duplication variants in autism spectrum disorder using long-read genome sequencing

    • Jesper Eisfeldt,
    • Edward J. Higginbotham,
    • Felix Lenner,
    • Jennifer Howe,
    • Bridget A. Fernandez,
    • Anna Lindstrand,
    • Stephen W. Scherer,
    • and Lars Feuk
    Genome Res. November 2024 34: 1763-1773; doi:10.1101/gr.279263.124

  • A national long-read sequencing study on chromosomal rearrangements uncovers hidden complexities

    • Jesper Eisfeldt,
    • Adam Ameur,
    • Felix Lenner,
    • Esmee Ten Berk de Boer,
    • Marlene Ek,
    • Josephine Wincent,
    • Raquel Vaz,
    • Jesper Ottosson,
    • Tord Jonson,
    • Sofie Ivarsson,
    • Sofia Thunström,
    • Alexandra Topa,
    • Simon Stenberg,
    • Anna Rohlin,
    • Anna Sandestig,
    • Margareta Nordling,
    • Pia Palmebäck,
    • Magnus Burstedt,
    • Frida Nordin,
    • Eva-Lena Stattin,
    • Maria Sobol,
    • Panagiotis Baliakas,
    • Marie-Louise Bondeson,
    • Ida Höijer,
    • Kristine Bilgrav Saether,
    • Lovisa Lovmar,
    • Hans Ehrencrona,
    • Malin Melin,
    • Lars Feuk,
    • and Anna Lindstrand
    Genome Res. November 2024 34: 1774-1784; doi:10.1101/gr.279510.124

  • Leveraging the T2T assembly to resolve rare and pathogenic inversions in reference genome gaps

    • Kristine Bilgrav Saether,
    • Jesper Eisfeldt,
    • Jesse D. Bengtsson,
    • Ming Yin Lun,
    • Christopher M. Grochowski,
    • Medhat Mahmoud,
    • Hsiao-Tuan Chao,
    • Jill A. Rosenfeld,
    • Pengfei Liu,
    • Marlene Ek,
    • Jakob Schuy,
    • Adam Ameur,
    • Hongzheng Dai,
    • Undiagnosed Diseases Network,
    • James Paul Hwang,
    • Fritz J. Sedlazeck,
    • Weimin Bi,
    • Ronit Marom,
    • Josephine Wincent,
    • Ann Nordgren,
    • Claudia M.B. Carvalho,
    • and Anna Lindstrand
    Genome Res. November 2024 34: 1785-1797; doi:10.1101/gr.279346.124

  • Independent expansion, selection, and hypervariability of the TBC1D3 gene family in humans

    • Xavi Guitart,
    • David Porubsky,
    • DongAhn Yoo,
    • Max L. Dougherty,
    • Philip C. Dishuck,
    • Katherine M. Munson,
    • Alexandra P. Lewis,
    • Kendra Hoekzema,
    • Jordan Knuth,
    • Stephen Chang,
    • Tomi Pastinen,
    • and Evan E. Eichler
    Genome Res. November 2024 34: 1798-1810; doi:10.1101/gr.279299.124

  • Unraveling the architecture of major histocompatibility complex class II haplotypes in rhesus macaques

    • Nanine de Groot,
    • Marit van der Wiel,
    • Ngoc Giang Le,
    • Natasja G. de Groot,
    • Jesse Bruijnesteijn,
    • and Ronald E. Bontrop
    Genome Res. November 2024 34: 1811-1824; doi:10.1101/gr.278968.124

  • Long-read DNA and cDNA sequencing identify cancer-predisposing deep intronic variation in tumor-suppressor genes

    • Suleyman Gulsuner,
    • Amal AbuRayyan,
    • Jessica B. Mandell,
    • Ming K. Lee,
    • Greta V. Bernier,
    • Barbara M. Norquist,
    • Sarah B. Pierce,
    • Mary-Claire King,
    • and Tom Walsh
    Genome Res. November 2024 34: 1825-1831; doi:10.1101/gr.279158.124

  • Long-read transcriptome sequencing of CLL and MDS patients uncovers molecular effects of SF3B1 mutations

    • Alicja Pacholewska,
    • Matthias Lienhard,
    • Mirko Brüggemann,
    • Heike Hänel,
    • Lorina Bilalli,
    • Anja Königs,
    • Felix Heß,
    • Kerstin Becker,
    • Karl Köhrer,
    • Jesko Kaiser,
    • Holger Gohlke,
    • Norbert Gattermann,
    • Michael Hallek,
    • Carmen D. Herling,
    • Julian König,
    • Christina Grimm,
    • Ralf Herwig,
    • Kathi Zarnack,
    • and Michal R. Schweiger
    Genome Res. November 2024 34: 1832-1848; doi:10.1101/gr.279327.124

  • Long-read RNA sequencing of archival tissues reveals novel genes and transcripts associated with clear cell renal cell carcinoma recurrence and immune evasion

    • Joshua Lee,
    • Elizabeth A. Snell,
    • Joanne Brown,
    • Charlotte E. Booth,
    • Rosamonde E. Banks,
    • Daniel J. Turner,
    • Naveen S. Vasudev,
    • and Dimitris Lagos
    Genome Res. November 2024 34: 1849-1864; doi:10.1101/gr.278801.123

  • Enhanced detection of RNA modifications and read mapping with high-accuracy nanopore RNA basecalling models

    • Gregor Diensthuber,
    • Leszek P. Pryszcz,
    • Laia Llovera,
    • Morghan C. Lucas,
    • Anna Delgado-Tejedor,
    • Sonia Cruciani,
    • Jean-Yves Roignant,
    • Oguzhan Begik,
    • and Eva Maria Novoa
    Genome Res. November 2024 34: 1865-1877; doi:10.1101/gr.278849.123

  • Long-read genome assembly of the insect model organism Tribolium castaneum reveals spread of satellite DNA in gene-rich regions by recurrent burst events

    • Marin Volarić,
    • Evelin Despot-Slade,
    • Damira Veseljak,
    • Brankica Mravinac,
    • and Nevenka Meštrović
    Genome Res. November 2024 34: 1878-1894; doi:10.1101/gr.279225.124

  • Genomic epidemiology of carbapenem-resistant Enterobacterales at a New York City hospital over a 10-year period reveals complex plasmid-clone dynamics and evidence for frequent horizontal transfer of blaKPC

    • Angela Gomez-Simmonds,
    • Medini K. Annavajhala,
    • Dwayne Seeram,
    • Todd W. Hokunson,
    • Heekuk Park,
    • and Anne-Catrin Uhlemann
    Genome Res. November 2024 34: 1895-1907; doi:10.1101/gr.279355.124

Methods

  • Telomere-to-telomere assembly by preserving contained reads

    • Sudhanva Shyam Kamath,
    • Mehak Bindra,
    • Debnath Pal,
    • and Chirag Jain
    Genome Res. November 2024 34: 1908-1918; doi:10.1101/gr.279311.124

  • Gapless assembly of complete human and plant chromosomes using only nanopore sequencing

    • Sergey Koren,
    • Zhigui Bao,
    • Andrea Guarracino,
    • Shujun Ou,
    • Sara Goodwin,
    • Katharine M. Jenike,
    • Julian Lucas,
    • Brandy McNulty,
    • Jimin Park,
    • Mikko Rautiainen,
    • Arang Rhie,
    • Dick Roelofs,
    • Harrie Schneiders,
    • Ilse Vrijenhoek,
    • Koen Nijbroek,
    • Olle Nordesjo,
    • Sergey Nurk,
    • Mike Vella,
    • Katherine R. Lawrence,
    • Doreen Ware,
    • Michael C. Schatz,
    • Erik Garrison,
    • Sanwen Huang,
    • William Richard McCombie,
    • Karen H. Miga,
    • Alexander H.J. Wittenberg,
    • and Adam M. Phillippy
    Genome Res. November 2024 34: 1919-1930; doi:10.1101/gr.279334.124

  • Full-resolution HLA and KIR gene annotations for human genome assemblies

    • Ying Zhou,
    • Li Song,
    • and Heng Li
    Genome Res. November 2024 34: 1931-1941; doi:10.1101/gr.278985.124

  • Characterizing tandem repeat complexities across long-read sequencing platforms with TREAT and otter

    • Niccoló Tesi,
    • Alex Salazar,
    • Yaran Zhang,
    • Sven van der Lee,
    • Marc Hulsman,
    • Lydian Knoop,
    • Sanduni Wijesekera,
    • Jana Krizova,
    • Anne-Fleur Schneider,
    • Maartje Pennings,
    • Kristel Sleegers,
    • Erik-Jan Kamsteeg,
    • Marcel Reinders,
    • and Henne Holstege
    Genome Res. November 2024 34: 1942-1953; doi:10.1101/gr.279351.124

  • Measuring X-Chromosome inactivation skew for X-linked diseases with adaptive nanopore sequencing

    • Sena A. Gocuk,
    • James Lancaster,
    • Shian Su,
    • Jasleen K. Jolly,
    • Thomas L. Edwards,
    • Doron G. Hickey,
    • Matthew E. Ritchie,
    • Marnie E. Blewitt,
    • Lauren N. Ayton,
    • and Quentin Gouil
    Genome Res. November 2024 34: 1954-1965; doi:10.1101/gr.279396.124

  • An optimized protocol for quality control of gene therapy vectors using nanopore direct RNA sequencing

    • Kathleen Zeglinski,
    • Christian Montellese,
    • Matthew E. Ritchie,
    • Monther Alhamdoosh,
    • Cédric Vonarburg,
    • Rory Bowden,
    • Monika Jordi,
    • Quentin Gouil,
    • Florian Aeschimann,
    • and Arthur Hsu
    Genome Res. November 2024 34: 1966-1975; doi:10.1101/gr.279405.124

  • DNA-m6A calling and integrated long-read epigenetic and genetic analysis with fibertools

    • Anupama Jha,
    • Stephanie C. Bohaczuk,
    • Yizi Mao,
    • Jane Ranchalis,
    • Benjamin J. Mallory,
    • Alan T. Min,
    • Morgan O. Hamm,
    • Elliott Swanson,
    • Danilo Dubocanin,
    • Connor Finkbeiner,
    • Tony Li,
    • Dale Whittington,
    • William Stafford Noble,
    • Andrew B. Stergachis,
    • and Mitchell R. Vollger
    Genome Res. November 2024 34: 1976-1986; doi:10.1101/gr.279095.124

  • Detecting m6A RNA modification from nanopore sequencing using a semisupervised learning framework

    • Haotian Teng,
    • Marcus Stoiber,
    • Ziv Bar-Joseph,
    • and Carl Kingsford
    Genome Res. November 2024 34: 1987-1999; doi:10.1101/gr.278960.124

  • Long-read subcellular fractionation and sequencing reveals the translational fate of full-length mRNA isoforms during neuronal differentiation

    • Alexander J. Ritter,
    • Jolene M. Draper,
    • Christopher Vollmers,
    • and Jeremy R. Sanford
    Genome Res. November 2024 34: 2000-2011; doi:10.1101/gr.279170.124

  • Long-read Ribo-STAMP simultaneously measures transcription and translation with isoform resolution

    • Pratibha Jagannatha,
    • Alexandra T. Tankka,
    • Daniel A. Lorenz,
    • Tao Yu,
    • Brian A. Yee,
    • Kristopher W. Brannan,
    • Cathy J. Zhou,
    • Jason G. Underwood,
    • and Gene W. Yeo
    Genome Res. November 2024 34: 2012-2024; doi:10.1101/gr.279176.124

  • Nanopore strand-specific mismatch enables de novo detection of bacterial DNA modifications

    • Xudong Liu,
    • Ying Ni,
    • Lianwei Ye,
    • Zhihao Guo,
    • Lu Tan,
    • Jun Li,
    • Mengsu Yang,
    • Sheng Chen,
    • and Runsheng Li
    Genome Res. November 2024 34: 2025-2038; doi:10.1101/gr.279012.124

  • Accurate bacterial outbreak tracing with Oxford Nanopore sequencing and reduction of methylation-induced errors

    • Mara Lohde,
    • Gabriel E. Wagner,
    • Johanna Dabernig-Heinz,
    • Adrian Viehweger,
    • Sascha D. Braun,
    • Stefan Monecke,
    • Celia Diezel,
    • Claudia Stein,
    • Mike Marquet,
    • Ralf Ehricht,
    • Mathias W. Pletz,
    • and Christian Brandt
    Genome Res. November 2024 34: 2039-2047; doi:10.1101/gr.278848.123

  • Factors impacting target-enriched long-read sequencing of resistomes and mobilomes

    • Ilya B. Slizovskiy,
    • Nathalie Bonin,
    • Jonathan E. Bravo,
    • Peter M. Ferm,
    • Jacob Singer,
    • Christina Boucher,
    • and Noelle R. Noyes
    Genome Res. November 2024 34: 2048-2060; doi:10.1101/gr.279226.124

Resources

  • High-coverage nanopore sequencing of samples from the 1000 Genomes Project to build a comprehensive catalog of human genetic variation

    • Jonas A. Gustafson,
    • Sophia B. Gibson,
    • Nikhita Damaraju,
    • Miranda P.G. Zalusky,
    • Kendra Hoekzema,
    • David Twesigomwe,
    • Lei Yang,
    • Anthony A. Snead,
    • Phillip A. Richmond,
    • Wouter De Coster,
    • Nathan D. Olson,
    • Andrea Guarracino,
    • Qiuhui Li,
    • Angela L. Miller,
    • Joy Goffena,
    • Zachary B. Anderson,
    • Sophie H.R. Storz,
    • Sydney A. Ward,
    • Maisha Sinha,
    • Claudia Gonzaga-Jauregui,
    • Wayne E. Clarke,
    • Anna O. Basile,
    • André Corvelo,
    • Catherine Reeves,
    • Adrienne Helland,
    • Rajeeva Lochan Musunuri,
    • Mahler Revsine,
    • Karynne E. Patterson,
    • Cate R. Paschal,
    • Christina Zakarian,
    • Sara Goodwin,
    • Tanner D. Jensen,
    • Esther Robb,
    • The 1000 Genomes ONT Sequencing Consortium,
    • University of Washington Center for Rare Disease Research (UW-CRDR),
    • Genomics Research to Elucidate the Genetics of Rare Diseases (GREGoR) Consortium,
    • William Richard McCombie,
    • Fritz J. Sedlazeck,
    • Justin M. Zook,
    • Stephen B. Montgomery,
    • Erik Garrison,
    • Mikhail Kolmogorov,
    • Michael C. Schatz,
    • Richard N. McLaughlin,
    • Jr.,
    • Harriet Dashnow,
    • Michael C. Zody,
    • Matt Loose,
    • Miten Jain,
    • and Evan E. Eichler
    • and Danny E. Miller
    Genome Res. November 2024 34: 2061-2073; doi:10.1101/gr.279273.124

  • Visualization and analysis of medically relevant tandem repeats in nanopore sequencing of control cohorts with pathSTR

    • Wouter De Coster,
    • Ida Höijer,
    • Inge Bruggeman,
    • Svenn D'Hert,
    • Malin Melin,
    • Adam Ameur,
    • and Rosa Rademakers
    Genome Res. November 2024 34: 2074-2080; doi:10.1101/gr.279265.124

  • Haplotype-resolved genome and population genomics of the threatened garden dormouse in Europe

    • Paige A. Byerly,
    • Alina von Thaden,
    • Evgeny Leushkin,
    • Leon Hilgers,
    • Shenglin Liu,
    • Sven Winter,
    • Tilman Schell,
    • Charlotte Gerheim,
    • Alexander Ben Hamadou,
    • Carola Greve,
    • Christian Betz,
    • Hanno J. Bolz,
    • Sven Büchner,
    • Johannes Lang,
    • Holger Meinig,
    • Evax Marie Famira-Parcsetich,
    • Sarah P. Stubbe,
    • Alice Mouton,
    • Sandro Bertolino,
    • Goedele Verbeylen,
    • Thomas Briner,
    • Lídia Freixas,
    • Lorenzo Vinciguerra,
    • Sarah A. Mueller,
    • Carsten Nowak,
    • and Michael Hiller
    Genome Res. November 2024 34: 2094-2107; doi:10.1101/gr.279066.124

  • Chromosome-level subgenome-aware de novo assembly provides insight into Saccharomyces bayanus genome divergence after hybridization

    • Cory Gardner,
    • Junhao Chen,
    • Christina Hadfield,
    • Zhaolian Lu,
    • David Debruin,
    • Yu Zhan,
    • Maureen J. Donlin,
    • Tae-Hyuk Ahn,
    • and Zhenguo Lin
    Genome Res. November 2024 34: 2133-2146; doi:10.1101/gr.279364.124

  • Construction and evaluation of a new rat reference genome assembly, GRCr8, from long reads and long-range scaffolding

    • Kai Li,
    • Melissa L. Smith,
    • J. Chris Blazier,
    • Kelli J. Kochan,
    • Jonathan M.D. Wood,
    • Kerstin Howe,
    • Anne E. Kwitek,
    • Melinda R. Dwinell,
    • Hao Chen,
    • Julia L. Ciosek,
    • Patrick Masterson,
    • Terence D. Murphy,
    • Theodore S. Kalbfleisch,
    • and Peter A. Doris
    Genome Res. November 2024 34: 2081-2093; doi:10.1101/gr.279292.124

  • Generation and analysis of a mouse multitissue genome annotation atlas

    • Matthew Adams,
    • and Christopher Vollmers
    Genome Res. November 2024 34: 2108-2117; doi:10.1101/gr.279217.124

  • Full-length RNA transcript sequencing traces brain isoform diversity in house mouse natural populations

    • Wenyu Zhang,
    • Anja Guenther,
    • Yuanxiao Gao,
    • Kristian Ullrich,
    • Bruno Huettel,
    • Aftab Ahmad,
    • Lei Duan,
    • Kaizong Wei,
    • and Diethard Tautz
    Genome Res. November 2024 34: 2118-2132; doi:10.1101/gr.279166.124
 

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