Wu Ting, Li Gang
Aging Institute, University of Pittsburgh, Pittsburgh, PA 15219, USA.
Department of Medicine, Xiangya School of Medicine, Central South University, Changsha, 410083, China.
Bio Protoc. 2022 Apr 20;12(8):e4397. doi: 10.21769/BioProtoc.4397.
Cells are the complex product of gene expression programs that involve the coordinated transcription of thousands of genes controlled by -regulatory elements (-REs). Therefore, identification of -REs is the key to decipher the mechanisms underlying the regulation of gene expression. Here, we describe a simple and time-effective protocol of fine-mapping -REs by using an electrophoresis mobility shift assay (EMSA)-based, , high-throughput (HTP) technique called regulatory element-sequencing (Reel-seq). Reel-seq can be applied to identify -REs at a high resolution and sensitivity over large genome regions, in a systematic and continuous manner. It can be used to prioritize candidate -REs as a complement to the existing approaches, such as massive parallel reporter assay (MPRA), chromatin immunoprecipitation DNA-sequencing (ChIP-seq), and the assay for transposase-accessible chromatin sequencing (ATAC-seq). NE: nuclear extract; NGS: next generation sequencing.
细胞是基因表达程序的复杂产物，这些程序涉及由调控元件（-REs）控制的数千个基因的协调转录。因此，鉴定-REs是破译基因表达调控机制的关键。在这里，我们描述了一种简单且省时的精细定位-REs的方案，该方案使用基于电泳迁移率变动分析（EMSA）的高通量（HTP）技术，称为调控元件测序（Reel-seq）。Reel-seq可用于在大基因组区域以系统且连续的方式高分辨率、高灵敏度地鉴定-REs。它可用于对候选-REs进行优先级排序，作为对现有方法（如大规模平行报告基因分析（MPRA）、染色质免疫沉淀DNA测序（ChIP-seq）和转座酶可及染色质测序分析（ATAC-seq））的补充。NE：核提取物；NGS：下一代测序。
