title: Measuring Arabidopsis chromatin accessibility using DNase I-polymerase chain reaction and DNase I-chip assays.
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.
authors.affiliation: Department of Biology and Zurich-Basel Plant Science Center, Eidgenössisch Technische Hochschule Zurich, CH-8092 Zurich, Switzerland.
authors.forename: Huan
authors.initials: H
authors.lastname: Shu
authors.forename: Wilhelm
authors.initials: W
authors.lastname: Gruissem
authors.forename: Lars
authors.initials: L
authors.lastname: Hennig
mesh_terms: D017360:Arabidopsis / Q000235:genetics; D002467:Cell Nucleus / Q000235:genetics; D002843:Chromatin / Q000235:genetics / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000235:genetics / Q000378:metabolism*; D018745:Genome, Plant; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D010539:Permeability; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D002843:Chromatin; D003850:Deoxyribonuclease I