用于结合/重塑分析的核小体体外组装

In Vitro Assembly of Nucleosomes for Binding/Remodeling Assays.

作者信息

Tran Kathleen Diep, Dutta Arnob

机构信息

University of Rhode Island, Center of Environmental and Life Sciences, Kingston, RI, USA.

Ocean State Research Institute (OSRI), Providence, RI, USA.

出版信息

Methods Mol Biol. 2025;2919:1-18. doi: 10.1007/978-1-0716-4486-7_1.

DOI:10.1007/978-1-0716-4486-7_1

PMID:40257554

Abstract

In order to study the functions and activities of chromatin remodeling enzymes in vitro, it is necessary to be able to reconstitute nucleosomes on DNA templates. In this chapter, we describe procedures for purification of histones from E. coli, formation of octamers, and reconstitution of nucleosomes, which can be further modified by chromatin modifiers. In addition, we describe methods to purify nucleosomes from human cells. Finally, we also describe assays to test binding and remodeling activities of chromatin remodelers.

摘要

为了在体外研究染色质重塑酶的功能和活性，必须能够在DNA模板上重建核小体。在本章中，我们描述了从大肠杆菌中纯化组蛋白、形成八聚体以及重建核小体的步骤，这些核小体可被染色质修饰剂进一步修饰。此外，我们还描述了从人类细胞中纯化核小体的方法。最后，我们还描述了测试染色质重塑因子结合和重塑活性的实验方法。