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Creating localized DNA double-strand breaks with microirradiation.


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Title: Creating localized DNA double-strand breaks with microirradiation.
Authors: Suzuki, Keiji / Yamauchi, Motohiro / Oka, Yasuyoshi / Suzuki, Masatoshi / Yamashita, Shunichi
Issue Date: Feb-2011
Publisher: Nature Publishing Group
Citation: Nature Protocols, 6(2), pp.134-139; 2011
Abstract: We describe a protocol for creating localized DNA double-strand breaks (DSBs) with minimal requirements that can be applied in cell biology and molecular biology. This protocol is based on the combination of 5-bromo-2'-deoxyuridine (BrdU) labeling and ultraviolet C (UVC) irradiation through porous membranes. Cells are labeled with 10 μM BrdU for 48-72 h, washed with Ca(2+)- and Mg(2+)-free PBS(-), covered by polycarbonate membranes with micropores and exposed to UVC light. With this protocol, localized DSBs are created within subnuclear areas, irrespective of the cell cycle phase. Recruitment of proteins involved in DNA repair, DNA damage response, chromatin remodeling and histone modifications can be visualized without any specialized equipment. The quality is the same as that obtained by laser microirradiation or by any other focal irradiation. DSBs become visible within 30 min of UVC irradiation.
Description: without figures
Keywords: DNA double strand breaks / micro-irradiation / DNA damage-induced foci / local damage / DNA damage response / DNA repair
URI: http://hdl.handle.net/10069/25685
ISSN: 17542189
DOI: 10.1038/nprot.2010.183
PubMed ID: 21293454
Rights: © 2011 Nature America, Inc. All rights reserved.
Type: Journal Article
Text Version: author
Appears in Collections:Articles in academic journal

Citable URI : http://hdl.handle.net/10069/25685

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