Measuring Arabidopsis chromatin accessibility using DNase I-polymerase chain reaction and DNase I-chip assays.
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Shu H, Gruissem W, Hennig L
Plant Physiol. 162 (4) 1794-1801 [2013-08-00; online 2013-06-05]
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.
PubMed 23739687
DOI 10.1104/pp.113.220400
Crossref 10.1104/pp.113.220400
pii: pp.113.220400
pmc: PMC3729761