Construction and application of a prokaryotic vector which expresses the protein that can be quickly purified by IMAC

A vector was constructed by inserting a pair of complementary oligo nucleotides encoding 6 histidine residues into the polylinker's upstream of the prokaryotic high expression vector pBV220. The resultant vector is named pBV222. Proteins expressed by this vector will have a 6-histidine tail as an affinity handle fused to their N-terminus and can be quickly purified by one-step immobilized metal affinity chromatography (IMAC). This plasmid was verified by restriction mapping and DNA sequencing. When GM-CSF and IL-2 cDNA were closed into pBV222, expressed proteins in the inclusion body showed the predicted molecular weight and biological activity. The expressed bacteria were dissolved in 6 mol/L guanidine.HCl and the supernatant was loaded directly to IMAC. IL-2 and GM-CSF fusion proteins were eluted by the pH gradient, and over 90% purity was achieved.