Fusion protein expression purification method

Abstract

The invention discloses a fusion protein expression purification method, which comprises the following experimental articles: 1ul of recombinant pET32a plasmids, BL21 (DE3) bacterial strains, 100ug / ml of penbritin, 0.5mM of IPTG (isopropyl-beta-d-thiogalactoside), PBS (Phosphate Buffer solution), 220rpm, 2M urea,50mM of Tris, 300mM of NaCl, 1mM of DTT (dithiothreitol), lysozyme, 0.2% of TritonX-100, 5mL of NI-NTA, EK (Enterokinase) enzyme, nickel agarose affinity chromatography and a SK3071 non-interference albumen concentration measurement kit. The fusion protein expression purification method comprises the following experiment process: the prokaryotic expression of fusion protein, the purification of the fusion protein and the analysis of the purified fusion protein. According to the fusion protein expression purification method, (50mM of imidazole) eluant which contains target protein is dialyzed into 1M urea, the 1M urea is dialyzed at a temperature of 4DEG C overnight, the EK enzyme is added into a sample subjected to dialysis overnight, protein subjected to enzyme digestion is subjected to the nickel agarose affinity chromatography to remove a label and the EK enzyme, and the purity of the purified fusion protein is detected and analyzed through SDS-PAGE (Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis) electrophoresis and gray analysis. Therefore, compared with a non-fusion expression carrier, the purified fusion protein has the advantages of high expression quantity, simple foreign protein separation and purification, high recovery rate, simple purification and purification technology and low cost.

Description

technical field [0001] The invention relates to the technical field of fusion protein expression and purification methods, in particular to a fusion protein expression and purification method. Background technique [0002] In the process of life science research and production of biological products, the use of expression vectors to prepare recombinant proteins is one of the most important technologies. Obtaining a large amount of active recombinant proteins is a necessary condition for the research and production of biological products. Constructing an effective expression vector is a basic requirement for expressing a target gene, and it is also an important factor affecting gene expression level and protein activity. Generally speaking, although the non-fusion expression vector can better maintain the primary structure of the foreign protein, its expression level is low, the separation and purification of the foreign protein is difficult, the recovery rate is low, the sep...

AI Technical Summary

Problems solved by technology

Generally speaking, although the non-fusion expression vector can better maintain the primary structure of the foreign protein, its expression level is low, the separation and purification of the foreign protein is difficult, the recovery rate is low, the separation and purification process is complicated, and large-scale industrial Higher cost of production

Images