Genome-Wide Mapping and Microscopy Visualization of Protein-DNA Interactions by pA-DamID
| Autor: | van Schaik, Tom, Manzo, Stefano G., van Steensel, Bas |
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| Přispěvatelé: | Cell biology |
| Jazyk: | angličtina |
| Rok vydání: | 2022 |
| Předmět: | |
| Zdroj: | Methods in Molecular Biology Methods in Molecular Biology-Chromatin Methods in Molecular Biology ISBN: 9781071621394 Methods in Molecular Biology, 2458, 215-229 |
| ISSN: | 1064-3745 1940-6029 |
| Popis: | Several methods have been developed to map protein–DNA interactions genome-wide in the last decades. Protein A-DamID (pA-DamID) is a recent addition to this list with distinct advantages. pA-DamID relies on antibody-based targeting of the bacterial Dam enzyme, resulting in adenine methylation of DNA in contact with the protein of interest. This m6A can then be visualized by microscopy, or mapped genome-wide. The main advantages of pA-DamID are an easy and direct visualization of DNA that is in contact with the protein of interest, unbiased mapping of protein–DNA interactions, and the possibility to select specific subpopulations of cells by flow cytometry before further sample processing. pA-DamID is particularly suited to study proteins that form large chromatin domains or that are part of distinct nuclear structures such as the nuclear lamina. This chapter describes the pA-DamID procedure from cell harvesting to the preparation of microscopy slides and high-throughput sequencing libraries. |
| Databáze: | OpenAIRE |
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