Suppr超能文献

蛋白质的一维十二烷基硫酸钠凝胶电泳。

One-dimensional SDS gel electrophoresis of proteins.

作者信息

Gallagher Sean R

机构信息

UVP, Inc., Upland, California, USA.

出版信息

Curr Protoc Cell Biol. 2007 Dec;Chapter 6:Unit 6.1. doi: 10.1002/0471143030.cb0601s37.

Abstract

Electrophoresis is used to separate complex mixtures of proteins (e.g., from cells, subcellular fractions, column fractions, or immunoprecipitates), to investigate subunit compositions, and to verify homogeneity of protein samples. It can also serve to purify proteins for use in further applications. In polyacrylamide gel electrophoresis, proteins migrate in response to an electrical field through pores in a polyacrylamide gel matrix; pore size decreases with increasing acrylamide concentration. The combination of pore size and protein charge, size, and shape determines the migration rate of the protein. In this unit, the standard Laemmli method is described for discontinuous gel electrophoresis under denaturing conditions, that is, in the presence of sodium dodecyl sulfate (SDS). Both full-size and minigel formats are detailed. Several modifications are provided for specific applications. For separation of peptides and small proteins, the standard buffers are replaced with either a Tris-tricine buffer system or a modified Tris buffer in the absence of urea. Continuous SDS-PAGE is a simplified method in which the same buffer is used for both the gel and the electrode solutions and the stacking gel is omitted. Other protocols cover the preparation and use of ultrathin gels and gradient gels, and the simultaneous preparation of multiple gels.

摘要

电泳用于分离蛋白质的复杂混合物(例如,从细胞、亚细胞组分、柱层析组分或免疫沉淀产物中分离),研究亚基组成,并验证蛋白质样品的纯度。它还可用于纯化蛋白质以用于进一步的应用。在聚丙烯酰胺凝胶电泳中,蛋白质在电场作用下通过聚丙烯酰胺凝胶基质中的孔隙迁移;孔隙大小随丙烯酰胺浓度的增加而减小。孔隙大小与蛋白质的电荷、大小和形状共同决定了蛋白质的迁移速率。在本单元中,将描述在变性条件下(即在十二烷基硫酸钠(SDS)存在的情况下)进行不连续凝胶电泳的标准Laemmli方法。详细介绍了全尺寸凝胶和迷你凝胶两种形式。针对特定应用提供了几种改进方法。对于肽和小蛋白质的分离,标准缓冲液可替换为Tris-三羟甲基氨基甲烷缓冲系统或不含尿素的改良Tris缓冲液。连续SDS-PAGE是一种简化方法,其中凝胶和电极溶液使用相同的缓冲液,并且省略了堆积凝胶。其他方案涵盖超薄凝胶和梯度凝胶的制备与使用,以及多种凝胶的同时制备。

文献AI研究员

20分钟写一篇综述,助力文献阅读效率提升50倍。

立即体验

用中文搜PubMed

大模型驱动的PubMed中文搜索引擎

马上搜索

文档翻译

学术文献翻译模型,支持多种主流文档格式。

立即体验