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Author: Ricke, R
Author: Bielinsky, A
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Methods Article

Easy detection of chromatin binding proteins by the histone association assay

Robin Ricke1 and Anja-Katrin Bielinsky1*

1 Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota. Minneapolis, MN 55455. USA.

* To whom correspondence should be addressed: Anja-Katrin Bielinsky, Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota. 321 Church Street SE, 6-155 Jackson Hall, Minneapolis, MN 55455. USA. Phone: 612-624-2469. Fax: 612-625-2163. Email: bielinsk@cbs.umn.edu

Biol. Proced. Online 2005;7:60-69. doi:10.1251/bpo106
Submitted: March 22, 2005; Accepted: April 14, 2005; Published: May 09, 2005.

Indexing terms: Chromatin; Cross-linking Reagents; Formaldehyde; Histones; Immunoprecipitation.

Abbreviations: ChIP, Chromatin Immunoprecipitation; HAA, Histone Association Assay; Mcm10, Minichromosome Maintenance Protein 10; Orc2, Origin Recognition Complex protein 2; WCE, Whole Cell Extract.


Figure 3 Enlarged

Fig. 3:

Efficiently crosslinked chromatin is not a suitable substrate for DNase I or MNase. ABy025 cells (MCM10-9MYC) were either crosslinked with formaldehyde (+) and quenched with glycine after 15 minutes or were left untreated (-). Formaldehyde crosslinked samples were sonicated to solubilize chromatin, while untreated samples were not. The extracts were treated with 300 U DNase I and 300 U MNase I for 30 minutes at either room temperature (RT) or 37°C. DNA was purified and visualized by ethidium bromide staining.

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