Chang Jennifer, Vancura Ales
Department of Cell Biology, New York University School of Medicine, New York, NY, USA.
Methods Mol Biol. 2012;809:321-33. doi: 10.1007/978-1-61779-376-9_22.
Chromatin remodeling is a key mode of transcriptional regulation, and studying the nucleosome positioning at promoters is an important means to understand how genes are regulated. Nucleosome scanning is a convenient method to study nucleosome positioning. Yeast cells are converted to spheroplasts and nuclei are isolated. The nuclei are then digested by micrococcal nuclease to yield mononucleosome-sized DNA. Using a set of overlapping primers that cover the entire promoter, quantitative real-time PCR is performed using the mononucleosome DNA as the template. The nucleosome enrichment for each primer is calculated to yield a map of nucleosome occupancy across the promoter.
染色质重塑是转录调控的关键模式，研究启动子处的核小体定位是理解基因如何被调控的重要手段。核小体扫描是研究核小体定位的一种便捷方法。将酵母细胞转化为原生质体并分离细胞核。然后用微球菌核酸酶消化细胞核以产生单核小体大小的DNA。使用一组覆盖整个启动子的重叠引物，以单核小体DNA为模板进行定量实时PCR。计算每个引物的核小体富集度，以生成启动子上核小体占据情况的图谱。
