Chapter title |
Chromosome Architecture
|
---|---|
Chapter number | 10 |
Book title |
Chromosome Architecture
|
Published in |
Methods in molecular biology, January 2016
|
DOI | 10.1007/978-1-4939-3631-1_10 |
Pubmed ID | |
Book ISBNs |
978-1-4939-3629-8, 978-1-4939-3631-1
|
Authors |
Miron, Ezequiel, Innocent, Cassandravictoria, Heyde, Sophia, Schermelleh, Lothar, Ezequiel Miron, Cassandravictoria Innocent, Sophia Heyde, Lothar Schermelleh |
Editors |
Mark C. Leake |
Abstract |
Recent advances in super-resolution microscopy enable the study of subchromosomal chromatin organization in single cells with unprecedented detail. Here we describe refined methods for pulse-chase replication labeling of individual chromosome territories (CTs) and replication domain units in mammalian cell nuclei, with specific focus on their application to three-dimensional structured illumination microscopy (3D-SIM). We provide detailed protocols for highly efficient electroporation-based delivery or scratch loading of cell impermeable fluorescent nucleotides for live cell studies. Furthermore we describe the application of (2'S)-2'-deoxy-2'-fluoro-5-ethynyluridine (F-ara-EdU) for the in situ detection of segregated chromosome territories with minimized cytotoxic side effects. |
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