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Plant Meiosis pp 381–402Cite as

In Planta Delivery of Chemical Compounds into Barley Meiocytes: EdU as Compound Example

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

Abstract

Here, we describe a protocol for in planta delivery of chemical compounds into meiocytes of different barley genotypes not impacting plant fertility allowing to harvest seeds from treated plants. Compound uptake into meiocytes is assessed by determining 5-ethynyl-2′-deoxyuridine (EdU) incorporation. Similar to EdU, other compounds being soluble in an aqueous solution can be delivered in planta before/during meiosis to decipher their impact on meiosis and meiotic recombination.

We give practical advice on how to deliver EdU as compound example (delivery via injection or needle and thread, addition of detergents or surfactants to increase compound uptake), how in planta compound delivery can be established for your plant material under specific growing conditions, how to generate and characterize barley hybrid plants, and how to conduct a meiotic cytological study of (treated) barley plants.

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References

  1. Mercier R, Mézard C, Jenczewski E, Macaisne N, Grelon M (2015) The molecular biology of meiosis in plants. Annu Rev Plant Biol 66:297–327. https://doi.org/10.1146/annurev-arplant-050213-035923

    Article  CAS  PubMed  Google Scholar 

  2. Wang Y, Copenhaver GP (2018) Meiotic recombination: mixing it up in plants. Annu Rev Plant Biol 69:577–609. https://doi.org/10.1146/annurev-arplant-042817-040431

    Article  CAS  PubMed  Google Scholar 

  3. Choulet F, Alberti A, Theil S, Glover N, Barbe V, Daron J et al (2014) Structural and functional partitioning of bread wheat chromosome 3B. Science 345:1249721. https://doi.org/10.1126/science.1249721

    Article  CAS  PubMed  Google Scholar 

  4. Phillips D, Wnetrzak J, Nibau C, Barakate A, Ramsay L, Wright F et al (2013) Quantitative high resolution mapping of HvMLH3 foci in barley pachytene nuclei reveals a strong distal bias and weak interference. J Exp Bot 64:2139–2154. https://doi.org/10.1093/jxb/ert079

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Fernandes JB, Séguéla-Arnaud M, Larchevêque C, Lloyd AH, Mercier R (2018) Unleashing meiotic crossovers in hybrid plants. Proc Natl Acad Sci U S A 115:2431–2436. https://doi.org/10.1073/pnas.1713078114

    Article  CAS  PubMed  Google Scholar 

  6. Ziolkowski PA, Underwood CJ, Lambing C, Martinez-Garcia M, Lawrence EJ, Ziolkowska L et al (2017) Natural variation and dosage of the HEI10 meiotic E3 ligase control Arabidopsis crossover recombination. Genes Dev 31:306–317. https://doi.org/10.1101/gad.295501.116

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Choi K, Zhao X, Tock AJ, Lambing C, Underwood CJ, Hardcastle TJ et al (2018) Nucleosomes and DNA methylation shape meiotic DSB frequency in Arabidopsis thaliana transposons and gene regulatory regions. Genome Res 28:532–546. https://doi.org/10.1101/gr.225599.117

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Underwood CJ, Choi K, Lambing C, Zhao X, Serra H, Borges F et al (2018) Epigenetic activation of meiotic recombination near Arabidopsis thaliana centromeres via loss of H3K9me2 and non-CG DNA methylation. Genome Res 28:519–531. https://doi.org/10.1101/gr.227116.117

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Yelina N, Diaz P, Lambing C, Henderson IR (2015) Epigenetic control of meiotic recombination in plants. Sci China Life Sci 58:223–231. https://doi.org/10.1007/s11427-015-4811-x

    Article  CAS  PubMed  Google Scholar 

  10. 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

    Article  PubMed  PubMed Central  Google Scholar 

  11. Fuchs LK, Jenkins G, Phillips DW (2018) Anthropogenic impacts on meiosis in plants. Front Plant Sci 9:1429. https://doi.org/10.3389/fpls.2018.01429

    Article  PubMed  PubMed Central  Google Scholar 

  12. Pecinka A, Liu CH (2014) Drugs for plant chromosome and chromatin research. Cytogenet Genome Res 143:51–59. https://doi.org/10.1159/000360774

    Article  CAS  PubMed  Google Scholar 

  13. Zhang H, Wang B, Duan CG, Zhu JK (2013) Chemical probes in plant epigenetics studies. Plant Signal Behav 8:e25364. https://doi.org/10.4161/psb.25364

    Article  PubMed  PubMed Central  Google Scholar 

  14. Perrella G, Consiglio MF, Aiese-Cigliano R, Cremona G, Sanchez-Moran E, Barra L et al (2010) Histone hyperacetylation affects meiotic recombination and chromosome segregation in Arabidopsis. Plant J 62:796–806. https://doi.org/10.1111/j.1365-313X.2010.04191.x

    Article  CAS  PubMed  Google Scholar 

  15. Griffing B, Langbridge J (1963) Factors affecting crossing over in the tomato. Aust J Biol Sci 16:826–837

    Article  CAS  Google Scholar 

  16. Knight E, Greer E, Draeger T, Thole V, Reader S, Shaw P et al (2010) Inducing chromosome pairing through premature condensation: analysis of wheat interspecific hybrids. Funct Integr Genomics 10:603–608. https://doi.org/10.1007/s10142-010-0185-0

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Colas I, Macaulay M, Higgins JD, Phillips D, Barakate A, Posch M et al (2016) A spontaneous mutation in MutL-Homolog 3 (HvMLH3) affects synapsis and crossover resolution in the barley desynaptic mutant des10. New Phytol 212:693–707. https://doi.org/10.1111/nph.14061

    Article  CAS  PubMed  Google Scholar 

  18. Higgins JD, Perry RM, Barakate A, Ramsay L, Waugh R, Halpin C et al (2012) Spatiotemporal asymmetry of the meiotic program underlies the predominantly distal distribution of meiotic crossovers in barley. Plant Cell 24:4096–4109. https://doi.org/10.1105/tpc.112.102483

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Sinha RP, Helgason SB (1969) The action of actinomycin D and diepoxybutane on recombination of two closely linked loci in Hordeum. Can J Genet Cytol 11:745–751

    Article  CAS  Google Scholar 

  20. Zadoks JC, Chang TT, Konzak CF (1974) A decimal code for the growth stages of cereals. Weed Res 14:415–421

    Article  Google Scholar 

  21. Gómez JF, Wilson ZA (2012) Non-destructive staging of barley reproductive development for molecular analysis based upon external morphology. J Exp Bot 63:4085–4094. https://doi.org/10.1093/jxb/ers092

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Tottman D, Makepeace R, Borad H (1979) An explanation of the decimal code for the growth stages of cereals, with illustrations. Ann Appl Biol 93:221–234

    Article  Google Scholar 

  23. Waddington S, Cartwright P, Wall P (1983) A quantitative scale of spike initial and pistil development in barley and wheat. Ann Bot 51:119–130

    Article  Google Scholar 

  24. Tracy SR, Gómez JF, Sturrock CJ, Wilson ZA, Ferguson AC (2017) Non-destructive determination of floral staging in cereals using X-ray micro computed tomography (μCT). Plant Methods 13:9. https://doi.org/10.1186/s13007-017-0162-x

    Article  PubMed  PubMed Central  Google Scholar 

  25. Arrieta M, Colas I, Macaulay M, Robbie Waugh R, Luke Ramsay L (2019) A modular tray growth system for barley. In: Plant meiosis: methods and protocols. Humana Press, Totowa, New Jersey

    Google Scholar 

  26. Bennett MD, Finch RA (1971) Duration of meiosis in barley. Genet Res 17:209–214

    Article  Google Scholar 

  27. Bennett MD, Finch RA, Smith JB, Rao MK (1973) The time and duration of female meiosis in wheat, rye and barley. Proc R Soc Lond B 183:301–319

    Article  Google Scholar 

  28. Finch RA, Bennett MD (1972) The duration of meiosis in diploid and autotetraploid barley. Can J Genet Cytol 14:507–515

    Article  Google Scholar 

  29. Dukowic-Schulze S, Garcia N, Shunmugam ASK, Kagale S, Chen C (2019) Isolating male meiocytes from maize and wheat for “-omics” analyses. In: Plant meiosis: methods and protocols. Humana Press, Totowa, New Jersey

    Google Scholar 

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Acknowledgments

We would like to thank the gardeners team from the IPK Gatersleben for their excellent support with plant cultivation, Jana Lorenz for excellent technical assistance, all members of the Meiosis team at the IPK for fruitful discussions, as well as the German Federal Ministry of Education and Research (BMBF—031B0188), Bayer CropScience AG (Grants4Traits 2016-1-18), the Marie-Curie “MEICOM” network H2020 ITN-765212, and the IPK Gatersleben for financial support.

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Authors and Affiliations

  1. Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Stadt Seeland, Germany

    Yun-Jae Ahn, Maria Cuacos, Judith Kappermann, Andreas Houben & Stefan Heckmann

  2. Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) OT Gatersleben, Stadt Seeland, Germany

    Mohammad A. Ayoub

Authors
  1. Yun-Jae Ahn
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  2. Maria Cuacos
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  3. Mohammad A. Ayoub
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  4. Judith Kappermann
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  5. Andreas Houben
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  6. Stefan Heckmann
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Corresponding author

Correspondence to Stefan Heckmann .

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Editors and Affiliations

  1. Departamento de Genética, Fisiología y Microbiología, Facultad de Biología, Universidad Complutense de Madrid, Madrid, Spain

    Mónica Pradillo

  2. Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) OT Gatersleben, Stadt Seeland, Germany

    Stefan Heckmann

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Ahn, YJ., Cuacos, M., Ayoub, M.A., Kappermann, J., Houben, A., Heckmann, S. (2020). In Planta Delivery of Chemical Compounds into Barley Meiocytes: EdU as Compound Example. 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_27

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  • DOI: https://doi.org/10.1007/978-1-4939-9818-0_27

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9817-3

  • Online ISBN: 978-1-4939-9818-0

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