APPLICATIONS OF GFP IN RESEARCH

Applications of GFP in Research

Abstract

The paper describes about the use of GFP protein in research for different purposes. It describes the summary of three relevant research papers and then analyzes its coding techniques. It also describes about the various aspects of the research studies and suggests alternatives for the GFP research.

Applications of GFP in Research

Introduction

The GFP protein consists of 238 amino acids that produces green fluorescence. It refers to the of jellyfish protein Aequorea victoria. In molecular biology, its gene is used for expression. It is also used for biosensors. This gene can be introduced into any organisms' genome by breeding, cell transformation or through a viral vector. This gene has been expressed in many Bacteria, and Fungi, plant, and mammalian cells. Osamu Shimomura, Martin Chalfie, and Roger Y. Tsien discovered it and were given Nobel Prize in 2008 for their discovery of the GFP.

In the 1960s, it was discovered and purified from Aequorea victoria . In this, GFP fluorescence is produced when aequorin reacts with Ca2+ ions and produces a green glow. In1992, Douglas Prasher discovered the nucleotide sequence of GFP. The Martin Chalfie lab expressed the sequence in E. coli cells. Later, the GFP was then folded at room temperature; this was sensitive to pH chloride and photosensitivity .

In 1996, Remington group in Science produced the crystals of GFP by S65T mutant. The Phillips group produced the wild-type GFP. These structures provided background for chromophore formation. This supports automated fluorescence microscopy systems of live-cells. For example, this can be used to label the spermatozoa of various species for identification, where GFP expression can be used as a marker. It is used to express the protein in specific cells. It allows detecting target cells. It is used to analyse brain circuitry and neuron membrane potential, to track AMPA receptors, influenza viruses' infection, and lentiviral viruses. It is useful for regenerative medicine and gene therapy.

Paper Summaries

The first paper “Expression and secretion of a CB4-1 scFv-GFP fusion protein by fission yeast” written by Naumann, Küttner & Bureik (2010) describes about the importance of fluorescent Fv (scFv) antibody fragments and its uses. It describes the usage of Yeasts as a primary hosts for producing these proteins. This is used for prokaryotes and eukaryotes. They reported that Schizosaccharomyces pombe can be used as a primary host for production of GFP. They had compared it with previous expression methods. They made a plasmid for anti-p24 and CB4-1 scFv fusion. They explained scFv-GFP fusion to the N-terminal of the Cpy1. They explained that the Western blot analysis provided accurate results that scFv-GFP was secreted and provided 5 mg/L of supernatants. The fluorescence analysis of supernatants provided evidence that GFP of scFv-GFP protein was functional after its secretion. They suggested that S. pombe can be used as a host for antibody fragment production for secretion of proteins (Naumann, Küttner & Bureik, 2010, pp. 80).

The second paper “Genotoxic evaluation of the insecticide endosulfan based on the induced GADD153-GFP reporter gene expression” written by ...