DNase I Chromatin Accessibility Analysis.

ABSTRACT

Chromatin consists of compacted DNA in complex with proteins and contributes to the organization of DNA and its stability. Furthermore, chromatin plays key roles in regulating cellular processes such as DNA replication, transcription, DNA repair, and mitosis. Chromatin assumes more compact (inaccessible) or decondensed (accessible) conformations depending on the function that is being supported in the genome, either locally or globally. The activity of nucleases has been used previously to assess the accessibility of specific genomic regions in vitro, such as origins of replication at varying points in the cell cycle. Here, we provide an assay to determine the accessibility of specific human genomic regions (example used herein Lamin B2 origin of DNA replication) by measuring the effect of DNase I nuclease on qPCR signal from the studied site. This assay provides a powerful method to interrogate the molecular mechanisms that regulate chromatin accessibility, and how these processes affect various cellular functions involving the human genome that require manipulation of chromatin conformation.