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Flow Cytometric Quantification, Isolation, and Subsequent Epigenetic Analysis of Tetraploid Neurons

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Genomic Mosaicism in Neurons and Other Cell Types

Part of the book series: Neuromethods ((NM,volume 131))

Abstract

Different forms of genomic mosaicism can be detected in vertebrate neurons, including copy number variation, L1 transposition, and aneuploidy. In addition, some populations of vertebrate neurons can also show double the normal amount of DNA, a condition referred to as somatic tetraploidy. These neurons are generated during early stages of development, as they migrate to their adult locations in the adult nervous system, and constitute subpopulations of projection neurons. We have previously shown that neuronal tetraploidy can be characterized by flow cytometry using isolated cell nuclei from different mammalian and avian structures. In this chapter, we describe this procedure using a different model system: the rhombencephalic derivatives from adult zebrafish. In addition, tetraploid neuronal nuclei can be isolated by fluorescence-activated cell sorting and their genomic DNA used for further analyses, either used directly or after whole-genome amplification. Here we show as an example how to perform epigenetic analyses to characterize CpG methylation in differentially methylated regions controlling the Rasgrf1-imprinting domain in mice.

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Acknowledgments

We thank Gonzalo García de Polavieja and Aixa Morales for the zebrafish specimens used in this study. This research was supported by the Ministerio de Economía y Competitividad (grant numbers SAF2015-68488-R) and a R&D contract with Tetraneuron S.L.

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Author notes

  1. Noelia López-Sánchez and Iris Patiño-Parrado are contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular, Cellular and Developmental Neurobiology, Cajal Institute (IC-CSIC), Avda. Doctor Arce 37, 28002, Madrid, Spain

    Noelia López-Sánchez, Iris Patiño-Parrado & José María Frade

  2. Tetraneuron S.L., Calle Duque de Calabria 13, Pta. 12, 46005, Valencia, Spain

    Noelia López-Sánchez

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  1. Noelia López-Sánchez
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  2. Iris Patiño-Parrado
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  3. José María Frade
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Correspondence to José María Frade .

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Editors and Affiliations

  1. Department of Molecular, Cellular and Developmental Neurobiology, Cajal Institute (IC-CSIC), Madrid, Spain

    José María Frade

  2. Biological Studies and Laboratory of Genetics, The Salk Institute, La Jolla, California, USA

    Fred H. Gage

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López-Sánchez, N., Patiño-Parrado, I., Frade, J.M. (2017). Flow Cytometric Quantification, Isolation, and Subsequent Epigenetic Analysis of Tetraploid Neurons. In: Frade, J., Gage, F. (eds) Genomic Mosaicism in Neurons and Other Cell Types. Neuromethods, vol 131. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7280-7_4

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  • DOI: https://doi.org/10.1007/978-1-4939-7280-7_4

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