Skip to main content

Targeted Analysis of Chromatin Events (TACE)

  • Protocol
  • First Online:
Plant Meiosis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2061))

Abstract

Visualization of meiotic chromatin from pollen mother cells has become an essential technique to study meiosis in the model plant Arabidopsis thaliana. Here we present an advanced cytogenetic method that combines improved immunocytology with chromosome painting, thereby generating a tool to quantitatively analyze localization of proteins to any given genomic region. Proteins involved in different processes such as DNA double-strand break formation and recombinational repair can be visualized on meiotic chromatin with the additional feature of assessing their abundance at specific chromosomal locations.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 109.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 199.99
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

Institutional subscriptions

Similar content being viewed by others

References

  1. Baltes NJ, Gil-Humanes J, Voytas DF (2017) Genome engineering and agriculture: opportunities and challenges. Prog Mol Biol Transl Sci 149:1–26. https://doi.org/10.1016/bs.pmbts.2017.03.011

    Article  PubMed  Google Scholar 

  2. Lambing C, Heckmann S (2018) Tackling plant meiosis: from model research to crop improvement. Front Plant Sci 9(829). https://doi.org/10.3389/fpls.2018.00829

  3. Berchowitz LE, Copenhaver GP (2008) Fluorescent Arabidopsis tetrads: a visual assay for quickly developing large crossover and crossover interference data sets. Nat Protoc 3(1):41–50. https://doi.org/10.1038/nprot.2007.491

    Article  CAS  PubMed  Google Scholar 

  4. Melamed-Bessudo C, Yehuda E, Stuitje AR, Levy AA (2005) A new seed-based assay for meiotic recombination in Arabidopsis thaliana. Plant J 43(3):458–466. https://doi.org/10.1111/j.1365-313X.2005.02466.x

    Article  CAS  PubMed  Google Scholar 

  5. Rowan BA, Patel V, Weigel D, Schneeberger K (2015) Rapid and inexpensive whole-genome genotyping-by-sequencing for crossover localization and fine-scale genetic mapping. G3 (Bethesda) 5(3):385–398. https://doi.org/10.1534/g3.114.016501

    Article  CAS  Google Scholar 

  6. Albini SM, Jones GH, Wallace BM (1984) A method for preparing two-dimensional surface-spreads of synaptonemal complexes from plant meiocytes for light and electron microscopy. Exp Cell Res 152(1):280–285. doi:0014-4827(84)90255-6 [pii]

    Article  CAS  Google Scholar 

  7. Albini SM, Jones GH (1984) Synaptonemal complex-associated centromeres and recombination nodules in plant meiocytes prepared by an improved surface-spreading technique. Exp Cell Res 155(2):588–592

    Article  CAS  Google Scholar 

  8. Armstrong SJ, Caryl AP, Jones GH, Franklin FC (2002) Asy1, a protein required for meiotic chromosome synapsis, localizes to axis-associated chromatin in Arabidopsis and Brassica. J Cell Sci 115(Pt 18):3645–3655

    Article  CAS  Google Scholar 

  9. Kurzbauer MT, Uanschou C, Chen D, Schlogelhofer P (2012) The recombinases DMC1 and RAD51 are functionally and spatially separated during meiosis in Arabidopsis. Plant Cell 24(5):2058–2070. https://doi.org/10.1105/tpc.112.098459

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Chen C, Farmer AD, Langley RJ, Mudge J, Crow JA, May GD, Huntley J, Smith AG, Retzel EF (2010) Meiosis-specific gene discovery in plants: RNA-Seq applied to isolated Arabidopsis male meiocytes. BMC Plant Biol 10:280. https://doi.org/10.1186/1471-2229-10-280

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Chen C, Retzel EF (2013) Analyzing the meiotic Transcriptome using isolated Meiocytes of Arabidopsis thaliana. Methods Mol Biol 990:203–213. https://doi.org/10.1007/978-1-62703-333-6_20

    Article  CAS  PubMed  Google Scholar 

  12. Bauman JG, Wiegant J, Borst P, van Duijn P (1980) A new method for fluorescence microscopical localization of specific DNA sequences by in situ hybridization of fluorochrome labelled RNA. Exp Cell Res 128(2):485–490

    Article  CAS  Google Scholar 

  13. Silahtaroglu A, Pfundheller H, Koshkin A, Tommerup N, Kauppinen S (2004) LNA-modified oligonucleotides are highly efficient as FISH probes. Cytogenet Genome Res 107(1–2):32–37. https://doi.org/10.1159/000079569

    Article  CAS  PubMed  Google Scholar 

  14. Paulasova P, Pellestor F (2004) The peptide nucleic acids (PNAs): a new generation of probes for genetic and cytogenetic analyses. Ann Genet 47(4):349–358. https://doi.org/10.1016/j.anngen.2004.07.001

    Article  PubMed  Google Scholar 

  15. Levsky JM, Singer RH (2003) Fluorescence in situ hybridization: past, present and future. J Cell Sci 116(Pt 14):2833–2838. https://doi.org/10.1242/jcs.00633

    Article  CAS  PubMed  Google Scholar 

  16. Lysak MA, Fransz PF, Ali HB, Schubert I (2001) Chromosome painting in Arabidopsis thaliana. Plant J 28(6):689–697

    Article  CAS  Google Scholar 

  17. Ried T, Schrock E, Ning Y, Wienberg J (1998) Chromosome painting: a useful art. Hum Mol Genet 7(10):1619–1626

    Article  CAS  Google Scholar 

  18. Miller AL, Gow NA (1989) Correlation between root-generated ionic currents, pH, Fusicoccin, Indoleacetic acid, and growth of the primary root of Zea mays. Plant Physiol 89(4):1198–1206

    Article  CAS  Google Scholar 

  19. Smyth DR, Bowman JL, Meyerowitz EM (1990) Early flower development in Arabidopsis. Plant Cell 2(8):755–767. https://doi.org/10.1105/tpc.2.8.755

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Wanzenbock EM, Schofer C, Schweizer D, Bachmair A (1997) Ribosomal transcription units integrated via T-DNA transformation associate with the nucleolus and do not require upstream repeat sequences for activity in Arabidopsis thaliana. Plant J 11(5):1007–1016

    Article  CAS  Google Scholar 

Download references

Acknowledgments

We thank the European Union (FP7-ITN 560 606956), the University of Vienna (I031-B), and the Austrian Science Fund (SFB F3408; P18036) for funding (J.S., P.S., and M.T.K.). C.C. is supported by the National Science Foundation (IOS:1025881 and IOS:1546792) and the Grant-in-Aid program at the University of Minnesota. We furthermore thank Chris Franklin for providing the ASY1 antibody and Sue Armstrong, James Higgins, and Katja Schneider for very helpful technical demonstrations and discussions.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Marie-Therese Kurzbauer .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2020 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Sims, J., Chen, C., Schlögelhofer, P., Kurzbauer, MT. (2020). Targeted Analysis of Chromatin Events (TACE). In: Pradillo, M., Heckmann, S. (eds) Plant Meiosis. Methods in Molecular Biology, vol 2061. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9818-0_5

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-9818-0_5

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9817-3

  • Online ISBN: 978-1-4939-9818-0

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics