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Plant Physiol. 2013 Aug;162(4):1794-801. doi: 10.1104/pp.113.220400. Epub 2013 Jun 5.

Measuring Arabidopsis chromatin accessibility using DNase I-polymerase chain reaction and DNase I-chip assays.

Author information

1
Department of Biology and Zurich-Basel Plant Science Center, Eidgenössisch Technische Hochschule Zurich, CH-8092 Zurich, Switzerland.

Abstract

DNA accessibility is an important layer of regulation of DNA-dependent processes. Methods that measure DNA accessibility at local and genome-wide scales have facilitated a rapid increase in the knowledge of chromatin architecture in animal and yeast systems. In contrast, much less is known about chromatin organization in plants. We developed a robust DNase I-polymerase chain reaction (PCR) protocol for the model plant Arabidopsis (Arabidopsis thaliana). DNA accessibility is probed by digesting nuclei with a gradient of DNase I followed by locus-specific PCR. The reduction in PCR product formation along the gradient of increasing DNase I concentrations is used to determine the accessibility of the chromatin DNA. We explain a strategy to calculate the decay constant of such signal reduction as a function of increasing DNase I concentration. This allows describing DNA accessibility using a single variable: the decay constant. We also used the protocol together with AGRONOMICS1 DNA tiling microarrays to establish genome-wide DNase I sensitivity landscapes.

PMID:
23739687
PMCID:
PMC3729761
DOI:
10.1104/pp.113.220400
[Indexed for MEDLINE]
Free PMC Article

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