蓝色原生聚丙烯酰胺凝胶电泳(BN-PAGE)分析植物蛋白质低聚物

Q1 Agricultural and Biological Sciences Current protocols in plant biology Pub Date : 2020-04-06 DOI:10.1002/cppb.20107

Pratchaya Pramoj Na Ayutthaya, Derek Lundberg, Detlef Weigel, Lei Li

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引用次数: 11

摘要

蛋白质之间的相互作用，包括寡聚化，参与了许多细胞过程的调节。不幸的是，许多蛋白质在体内的表达水平非常低，这使得通过尺寸排除色谱法(也称为凝胶过滤)观察寡聚化具有挑战性。在这个方案中，我们提出了详细的步骤来执行蓝色天然聚丙烯酰胺凝胶电泳(BN-PAGE)，一种研究植物蛋白质低聚物的方法。本文描述了从转基因拟南芥中制备蛋白质样品，并运行BN-PAGE凝胶，然后进行直接免疫印迹或二维十二烷基硫化钠-聚丙烯酰胺凝胶电泳(2D SDS-PAGE)。该协议将有助于新的研究人员进行相关实验来分析植物中稳定的蛋白质相互作用，包括同源或异寡聚。©2020作者。

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Blue Native Polyacrylamide Gel Electrophoresis (BN-PAGE) for the Analysis of Protein Oligomers in Plants

Protein-protein interactions, including oligomerization, are involved in regulation of many cellular processes. Unfortunately, many proteins are expressed at a very low level in vivo, making it challenging to observe oligomerization by size-exclusion chromatography, also known as gel filtration. In this protocol, we present detailed steps to perform blue native polyacrylamide gel electrophoresis (BN-PAGE), a method to study protein oligomers in plants. The article describes protein sample preparation from transgenic Arabidopsis thaliana and running a BN-PAGE gel followed by direct western blotting or, alternatively, two-dimensional sodium dodecyl sulfide–polyacrylamide gel electrophoresis (2D SDS-PAGE). This protocol will be helpful for new researchers conducting related experiments to analyze stable protein interactions including homo- or hetero-oligomerization in plants. © 2020 The Authors.

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Current protocols in plant biology Agricultural and Biological Sciences-Plant Science

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期刊介绍： Sound and reproducible laboratory methods are the foundation of scientific discovery. Yet nuances that are critical for an experiment''s success are not captured in the primary literature but exist only as part of a lab''s oral tradition. Current Protocols in Plant Biology provides reproducible step-by-step instructions for protocols relevant to plant research. Furthermore, Current Protocols content is thoughtfully organized by topic for optimal usage and to maximize contextual knowledge. Quarterly issues allow Current Protocols in Plant Biology to constantly evolve to keep pace with the newest discoveries and developments. Current Protocols in Plant Biology is the comprehensive source for protocols in the multidisciplinary field of plant biology, providing an extensive range of protocols from basic to cutting edge. Coverage includes: Extraction and analysis of DNA, RNA, proteins Chromosome analysis Transcriptional analysis Protein expression Metabolites Plant enzymology Epigenetics Plant genetic transformation Mutagenesis Arabidopsis, Maize, Poplar, Rice, and Soybean, and more.