Skip to main content
SpringerLink
Log in
Book cover

Transposable Elements pp 151–171Cite as

Nanopore Sequencing to Identify Transposable Element Insertions and Their Epigenetic Modifications

Part of the Methods in Molecular Biology book series (MIMB,volume 2607)

Abstract

Over the past 20 years, high-throughput genomic assays have fundamentally changed how transposable elements (TEs) are studied. While short-read DNA sequencing has been at the heart of these efforts, novel technologies that generate longer reads are driving a shift in the field. Long-read sequencing now permits locus-specific approaches to locate individual TE insertions and understand their epigenetic and transcriptional regulation, while still profiling TE activity genome-wide. Here we provide detailed guidelines to implement Oxford Nanopore Technologies (ONT) sequencing to identify polymorphic TE insertions and profile TE epigenetic landscapes. Using human long interspersed element-1 (LINE-1, L1) as an example, we explain the procedures involved, including final visualization, and potential bottlenecks and pitfalls. ONT sequencing will be, in our view, a workhorse technology for the foreseeable future in the TE field.

Key words

  • Transposable elements
  • LINE-1
  • Long-read sequencing
  • Nanopore sequencing
  • Epigenetics
  • DNA methylation
  • Mapping
  • Sequencing
  • Polymorphism
  • Basecalling

This is a preview of subscription content, access via your institution.

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   169.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   159.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   219.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Lander ES, Linton LM, Birren B et al (2001) Initial sequencing and analysis of the human genome. Nature 409:860–921. https://doi.org/10.1038/35057062

    CrossRef  CAS  PubMed  Google Scholar 

  2. Cost G, Feng Q, Jacquier A, Boeke JD (2002) Human L1 element target-primed reverse transcription in vitro. EMBO J 21:5899–5910. https://doi.org/10.1093/emboj/cdf592

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  3. Symer DE, Connelly C, Szak ST et al (2002) Human L1 retrotransposition is associated with genetic instability in vivo. Cell 110:327–338. https://doi.org/10.1016/S0092-8674(02)00839-5

    CrossRef  CAS  PubMed  Google Scholar 

  4. Castro-Diaz N, Ecco G, Coluccio A et al (2014) Evolutionally dynamic L1 regulation in embryonic stem cells. Genes Dev 28:1397–1409. https://doi.org/10.1101/gad.241661.114

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  5. de la Rica L, Deniz Ö, Cheng KCL et al (2016) TET-dependent regulation of retrotransposable elements in mouse embryonic stem cells. Genome Biol 17:234–234. https://doi.org/10.1186/s13059-016-1096-8

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  6. Fadloun A, Le Gras S, Jost B et al (2013) Chromatin signatures and retrotransposon profiling in mouse embryos reveal regulation of LINE-1 by RNA. Nat Struct Mol Biol 20:332–338. https://doi.org/10.1038/nsmb.2495

    CrossRef  CAS  PubMed  Google Scholar 

  7. Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A 74:5463–5467. https://doi.org/10.1073/pnas.74.12.5463

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  8. Churko JM, Mantalas GL, Snyder MP, Wu JC (2013) Overview of high throughput sequencing technologies to elucidate molecular pathways in cardiovascular diseases. Circ Res 112:1613–1623. https://doi.org/10.1161/CIRCRESAHA.113.300939

    CrossRef  CAS  PubMed  Google Scholar 

  9. Treangen TJ, Salzberg SL (2011) Repetitive DNA and next-generation sequencing: computational challenges and solutions. Nat Rev Genet 13:36–46. https://doi.org/10.1038/nrg3117

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  10. Lanciano S, Cristofari G (2020) Measuring and interpreting transposable element expression. Nat Rev Genet 21:721–736. https://doi.org/10.1038/s41576-020-0251-y

    CrossRef  CAS  PubMed  Google Scholar 

  11. Ewing AD (2015) Transposable element detection from whole genome sequence data. Mob DNA 6:1–9

    CrossRef  Google Scholar 

  12. Scott EC, Gardner EJ, Masood A et al (2016) A hot L1 retrotransposon evades somatic repression and initiates human colorectal cancer. Genome Res 26:745–755. https://doi.org/10.1101/gr.201814.115

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  13. Berrens RV, Yang A, Laumer CE et al (2021) Locus-specific expression of transposable elements in single cells with CELLO-seq. Nat Biotechnol 40:546. https://doi.org/10.1038/s41587-021-01093-1

    CrossRef  CAS  PubMed  Google Scholar 

  14. Faulkner GJ, Garcia-Perez JL (2017) L1 mosaicism in mammals: extent, effects, and evolution. Trends Genet 33:802–816

    CrossRef  CAS  PubMed  Google Scholar 

  15. Faulkner GJ, Billon V (2018) L1 retrotransposition in the soma: a field jumping ahead. Mob DNA 9:1–18

    CrossRef  Google Scholar 

  16. Sanchez-Luque FJ, Kempen MJHC, Gerdes P et al (2019) LINE-1 evasion of epigenetic repression in humans. Mol Cell 75:590–604.e12. https://doi.org/10.1016/j.molcel.2019.05.024

    CrossRef  CAS  PubMed  Google Scholar 

  17. Ewing AD, Smits N, Sanchez-Luque FJ et al (2020) Nanopore sequencing enables comprehensive transposable element epigenomic profiling. Mol Cell 80:915–928.e5. https://doi.org/10.1016/j.molcel.2020.10.024

    CrossRef  CAS  PubMed  Google Scholar 

  18. Zhou W, Emery SB, Flasch DA et al (2020) Identification and characterization of occult human-specific LINE-1 insertions using long-read sequencing technology. Nucleic Acids Res 48:1146–1163. https://doi.org/10.1093/nar/gkz1173

    CrossRef  CAS  PubMed  Google Scholar 

  19. Deininger P, Morales ME, White TB et al (2017) A comprehensive approach to expression of L1 loci. Nucleic Acids Res 45:e31

    CrossRef  PubMed  Google Scholar 

  20. Gouil Q, Keniry A (2019) Latest techniques to study DNA methylation. Essays Biochem 63:639–648. https://doi.org/10.1042/EBC20190027

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  21. Logsdon GA, Vollger MR, Eichler EE (2020) Long-read human genome sequencing and its applications. Nat Rev Genet 21:1–18

    CrossRef  Google Scholar 

  22. McDonald TL, Zhou W, Castro CP et al (2021) Cas9 targeted enrichment of mobile elements using nanopore sequencing. Nat Commun 12:3586. https://doi.org/10.1038/s41467-021-23918-y

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  23. Wang Y, Zhao Y, Bollas A et al (2021) Nanopore sequencing technology, bioinformatics and applications. Nat Biotechnol 39:1–18. https://doi.org/10.1038/s41587-021-01108-x

    CrossRef  CAS  Google Scholar 

  24. Siudeja K, van den Beek M, Riddiford N et al (2021) Unraveling the features of somatic transposition in the Drosophila intestine. EMBO J 40:e106388. https://doi.org/10.15252/embj.2020106388

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  25. Deamer D, Akeson M, Branton D (2016) Three decades of nanopore sequencing. Nat Biotechnol 34:518–524. https://doi.org/10.1038/nbt.3423

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  26. Shendure J, Balasubramanian S, Church GM et al (2017) DNA sequencing at 40: past, present and future. Nature 550:345–353. https://doi.org/10.1038/nature24286

    CrossRef  CAS  PubMed  Google Scholar 

  27. Smits N, Rasmussen J, Bodea GO et al (2021) No evidence of human genome integration of SARS-CoV-2 found by long-read DNA sequencing. Cell Rep 36:109530. https://doi.org/10.1016/j.celrep.2021.109530

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  28. Requirements PIIT PromethION P24/P48 IT requirements. https://community.nanoporetech.com/requirements_documents/promethion-it-reqs.pdf

  29. Miniconda – conda documentation. https://docs.conda.io/en/latest/miniconda.html. Accessed 4 Mar 2022

  30. Li H (2018) Minimap2: pairwise alignment for nucleotide sequences. Bioinformatics 34:3094–3100

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  31. Robinson JT, Thorvaldsdóttir H, Winckler W et al (2011) Integrative genomics viewer. Nat Biotechnol 29:24–26. https://doi.org/10.1038/nbt.1754

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  32. Yoder JA, Walsh CP, Bestor TH (1997) Cytosine methylation and the ecology of intragenomic parasites. Trends Genet 13:335–340

    CrossRef  CAS  PubMed  Google Scholar 

  33. Greenberg MVC, Bourc’his D (2019) The diverse roles of DNA methylation in mammalian development and disease. Nat Rev Mol Cell Biol 20:590–607. https://doi.org/10.1038/s41580-019-0159-6

    CrossRef  CAS  PubMed  Google Scholar 

  34. Liu Y, Rosikiewicz W, Pan Z et al (2021) DNA methylation-calling tools for Oxford Nanopore sequencing: a survey and human epigenome-wide evaluation. Genome Biol 22:1–33. https://doi.org/10.1186/s13059-021-02510-z

    CrossRef  CAS  Google Scholar 

  35. Gamaarachchi H, Lam CW, Jayatilaka G et al (2020) GPU accelerated adaptive banded event alignment for rapid comparative nanopore signal analysis. BMC Bioinformatics 21:343. https://doi.org/10.1186/s12859-020-03697-x

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  36. Simpson JT, Workman RE, Zuzarte PC et al (2017) Detecting DNA cytosine methylation using nanopore sequencing. Nat Methods 14:407–410. https://doi.org/10.1038/nmeth.4184

    CrossRef  CAS  PubMed  Google Scholar 

  37. Su S, Gouil Q, Blewitt ME et al (2021) NanoMethViz: an R/Bioconductor package for visualizing long-read methylation data. PLoS Comput Biol 17:e1009524. https://doi.org/10.1371/journal.pcbi.1009524

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  38. De Coster W, Stovner EB, Strazisar M (2020) Methplotlib: analysis of modified nucleotides from nanopore sequencing. Bioinformatics 36:3236–3238. https://doi.org/10.1093/bioinformatics/btaa093

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  39. Lee I, Razaghi R, Gilpatrick T et al (2020) Simultaneous profiling of chromatin accessibility and methylation on human cell lines with nanopore sequencing. Nat Methods 17:1191–1199. https://doi.org/10.1038/s41592-020-01000-7

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  40. Gamaarachchi H, Samarakoon H, Jenner SP et al (2022) Fast nanopore sequencing data analysis with SLOW5. Nat Biotechnol 40:1026–1029. https://doi.org/10.1038/s41587-021-01147-4

Download references

Author information

Authors and Affiliations

  1. Mater Research Institute, University of Queensland, Woolloongabba, QLD, Australia

    Nathan Smits & Geoffrey J. Faulkner

  2. Queensland Brain Institute, University of Queensland, Brisbane, QLD, Australia

    Geoffrey J. Faulkner

Authors
  1. Nathan Smits
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Geoffrey J. Faulkner
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Faculty of Medicine and Dentistry, Blizard Institute, Queen Mary University of London, London, UK

    Miguel R. Branco

  2. School of Biological and Behavioural Sciences, Queen Mary University of London, London, UK

    Alexandre de Mendoza Soler

Rights and permissions

Reprints and Permissions

Copyright information

© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Smits, N., Faulkner, G.J. (2023). Nanopore Sequencing to Identify Transposable Element Insertions and Their Epigenetic Modifications. In: Branco, M.R., de Mendoza Soler, A. (eds) Transposable Elements. Methods in Molecular Biology, vol 2607. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2883-6_9

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-2883-6_9

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2882-9

  • Online ISBN: 978-1-0716-2883-6

  • eBook Packages: Springer Protocols

search

Navigation