SDSPAGE实验报告.pdfVIP

SDS Lin Chengyu Bio 04 2010030007; Cooperator: Liu Yidi 1 Introduction 1.1 Background information SDS, which is abbreviated for sodium dodecyl sulphate-polyacrylamide gel electrophoresis, is the most widely used method for analyzing protein mixtures qualitatively. It is particularly useful for monitoring protein purification and, because the method is based on the separation of proteins according to size, the method can also be used to determine the relative molecular mass of proteins. This experiment applies SDSto rabbit muscle creatine kinase to examine its purity and molecular weight. 1.2 Major principles 1.2.1 SDS SDS is an anionic detergent. When protein molecules are treated with SDS, the detergent disrupts the secondary, tertiary, and quaternary structure to produce linear polypeptide chains coated with negatively charged SDS molecules. But sometimes disulfide bound also exist in the protein molecule to link different subunit, so mercaptoethanol assists in protein denaturation by reducing all disulfide bonds. The protein molecule opens up into a rod-shaped structure with a series of negatively charged SDS molecules alongside with a constant charge/mass ratio. 1.2.2 PAGE The electrophoretic mobility of the SDS-protein complexes is influenced primarily by molecular size: the larger molecules are retarded by molecular sieving effect of the gel, and the smaller molecules have greater mobility