利用荧光偏振法对AGO蛋白进行核酸结合分析

Nucleic Acid-Binding Assay of Argonaute Protein Using Fluorescence Polarization.

作者信息

Miyoshi Tomohiro

机构信息

Center for Transdisciplinary Research, Niigata University, 8050 Ikarashi 2-no-cho, Nishi-ku, Niigata, 950-2181, Japan.

出版信息

Methods Mol Biol. 2018;1680:123-129. doi: 10.1007/978-1-4939-7339-2_8.

DOI:10.1007/978-1-4939-7339-2_8

PMID:29030845

Abstract

Nucleic acid binding by the Argonaute protein is an important trigger step in the Argonaute-dependent gene silencing system. We established an in vitro method to detect the nucleic acid binding activity of the Argonaute protein by fluorescence polarization. In this chapter, we will describe the expression and purification of the prokaryotic (Rhodobacter sphaeroides) Argonaute protein, and the nucleic acid-binding analysis using a Fluorescence Polarization System (Beacon 2000).

摘要

在依赖于Argonaute的基因沉默系统中，Argonaute蛋白与核酸的结合是一个重要的触发步骤。我们建立了一种体外方法，通过荧光偏振来检测Argonaute蛋白的核酸结合活性。在本章中，我们将描述原核生物（球形红细菌）Argonaute蛋白的表达和纯化，以及使用荧光偏振系统（Beacon 2000）进行的核酸结合分析。