Quantitative characterization of specific genomic promoters using agarose gel electrophoresis

Abstract

Over the past decade a large number of studies have focused attention on the role of nucleosomes as negative and positive regulators of specific nuclear functions. Due to the lack of an analytical method to determine the higher order conformation of the nucleosomal arrays that encompass specific genetic loci (e.g., promoters, enhancers), research emphasis has mostly been centered on chromatin remodeling and histone posttranslational modifications. We have recently developed an agarose gel electrophoresis method that permits us to analyze the higher order structure of specific in vivo assembled chromatin fragments. After calibration using a well-defined in vitro system, we have been able to experimentally determine the size, shape, and conformational flexibility of the Mouse Mammary Tumor Virus long-terminal repeat promoter region in its repressed and activated states. These studies pave the way for widespread analyses of the higher order structure of specific, functionally important chromosomal loci, and in so doing enhance our understanding of the roles that the higher order structure of chromatin play in genome regulations.