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Imaging Replicative Domains in Ultrastructurally Preserved Chromatin by Electron Tomography
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Biologia
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JoVE Journal Biologia
Imaging Replicative Domains in Ultrastructurally Preserved Chromatin by Electron Tomography

Imaging Replicative Domains in Ultrastructurally Preserved Chromatin by Electron Tomography

DOI:
10.3791/62803-v

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14:56 min

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May 20, 2022

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, , , , , , , , ,
1Belozersky Institute of Physico-chemical Biology,Lomonosov Moscow State University, 2Faculty of Biology,Lomonosov Moscow State University, 3Faculty of Bioengineering and Bioinformatics,Lomonosov Moscow State University, 4National Research Center, Kurchatov Institute, 5V.I. Kulakov National Medical Research Center for Obstetrics, Gynecology, and Perinatology

Capítulos

  • 00:08Introduction
  • 00:35Cells Labeling and Fixation
  • 02:13Click-Reaction
  • 05:09Ag-Amplification
  • 06:28Gold Toning
  • 06:42ChromEM
  • 07:23Dehydration and Epon Embedding
  • 12:50Electron Tomography
  • 13:06Representative Results
  • 14:14Conclusions

Summary

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This protocol presents a technique for high-resolution mapping of replication sites in structurally preserved chromatin in situ that employs a combination of pre-embedding EdU-streptavidin-Nanogold labeling and ChromEMT.

Tags

Keywords: Electron TomographyReplicative DomainsChromatinEdU LabelingSilver EnhancementChromEM StainingGlutaraldehyde FixationPlasma Membrane PermeabilizationBiotin-azide ConjugationStreptavidin-Nanogold

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