Expression and purification of the Green Fluorescent Protein via immobilized metal ion affinity.Olga D. SieluzyckaSchool of Biological and Chemical Sciences (Fogg Building)Queen Mary University of London Mile End RoadLondon E14NS UKABSTRACT: The aim of the experiment is to express and purify recombinant His-tagged Green Fluorescent Protein (GFP) protein inEscherichia coli (E.coli) using metal affinity chromatography. GFP is an excellent trafficking tool in modern biochemistry. The protein was expressed in E.Coli and introduced to the required organism by a plasmid transformation. Plasmid containing the GFP was controlled by the T7 promoter had ampicillin resistance and had an N-terminal His6 tag fusion to facilitate the process of purification in the later stages of experiment. The expression of proteins was confirmed by the SDS gel by comparing the bands widths. The goal of experiment was not fully met.Introduction: The main objective of the experiment is to express and purify recombinant His tagged, Green Fluorescent Protein (GFP). GFP is a protein native to a jellyfish, Aequorea Victoriaand due to its remarkable fluorescence it has a vast amount of applications in modern biochemistry such as in vivo labelling and visualization of cells structures. Since the GFPs fluorescence disturbed by unfavorable conditions can recover in the correct pH and temperature it is incredibly useful in molecular biology and its ability to track “real time” changes in cells with a “naked eye” captured the attention of various researchers. (Wu et al., 2008) Another big advantage of using GFP over other molecular trackers is that it does not require any substrates or cofactors has a low toxicity towards examined cells and does not alter the localization of its fusion partner. (Kumar et al., 2016)