In recent years, along with the rapid development of genetic engineering technologies, a vast amount of valuable proteins have been expressed in Escherichia coli with high productivity. In order to produce large amounts of human dystrophin Dp71, a protein of unknown function, its coding sequence was cloned into the prokaryotic expression vector pMAL-c2X generating the gene fusion malE-Dp71. Screening of BL21, a protease-deficient strain, expressing the gene fusion malE-Dp71 was achieved by SDS-PAGE gel and Western blotting with an anti-Dp71 polyclonal antibody. The maltose-binding protein-Dp71 (MBP-Dp71) fusion protein, expressed in E. coli, was recovered into the soluble fraction of cell lysates using the sarkosyl detergent, and purified to homogeneity by affinity chromatography on amylose columns. The expression and purification procedures yielded approximately 98.5 μg of highly purified protein per 100 ml of bacterial culture. The expression system established in this study will provide the biological material needed for in vitro studies of Dp71.
|Number of pages||5|
|Journal||Annals of Microbiology|
|Publication status||Published - 11 Jul 2005|