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Genome-Wide Cell Type-Specific Mapping of In Vivo Chromatin Protein Binding Using an FLP-Inducible DamID System in Drosophila

  • Alexey V. Pindyurin
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1654)

Abstract

A thorough study of the genome-wide binding patterns of chromatin proteins is essential for understanding the regulatory mechanisms of genomic processes in eukaryotic nuclei, including DNA replication, transcription, and repair. The DNA adenine methyltransferase identification (DamID) method is a powerful tool to identify genomic binding sites of chromatin proteins. This method does not require fixation of cells and the use of specific antibodies, and has been used to generate genome-wide binding maps of more than a hundred different proteins in Drosophila tissue culture cells. Recent versions of inducible DamID allow performing cell type-specific profiling of chromatin proteins even in small samples of Drosophila tissues that contain heterogeneous cell types. Importantly, with these methods sorting of cells of interest or their nuclei is not necessary as genomic DNA isolated from the whole tissue can be used as an input. Here, I describe in detail an FLP-inducible DamID method, namely generation of suitable transgenic flies, activation of the Dam transgenes by the FLP recombinase, isolation of DNA from small amounts of dissected tissues, and subsequent identification of the DNA binding sites of the chromatin proteins.

Key words

Drosophila melanogaster Chromatin proteins Genomic binding sites DamID Cell type-specific profiling “Flp-Out” approach 

Notes

Acknowledgments

I thank the laboratory of Prof. B. van Steensel at the Netherlands Cancer Institute (Amsterdam, the Netherlands) for providing an excellent working environment, the NKI Genomics Core Facility for help with the development of the protocol for preparation of DNA samples for Illumina high-throughput sequencing; Anna A. Ogienko for technical support; Mario Amendola, Maurizio Gatti, and the members of Laboratory of Cell Division for critical reading of the manuscript and helpful suggestions. This work was supported by the grant of Russian Science Foundation no. 16-14-10288.

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Laboratory of Cell DivisionInstitute of Molecular and Cellular Biology SB RASNovosibirskRussia

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