World Library  
Flag as Inappropriate
Email this Article

Reporter gene

Article Id: WHEBN0000341038
Reproduction Date:

Title: Reporter gene  
Author: World Heritage Encyclopedia
Language: English
Subject: Two-hybrid screening, Bioreporter, Host-Cell Reactivation, Enhancer-FACS-seq, Vector (molecular biology)
Collection: Genetics, Molecular Biology
Publisher: World Heritage Encyclopedia
Publication
Date:
 

Reporter gene

A diagram of a how a reporter gene is used to study a regulatory sequence.

In selectable markers. Reporter genes are often used as an indication of whether a certain gene has been taken up by or expressed in the cell or organism population.

Contents

  • Common reporter genes 1
  • Transformation and transfection assays 2
  • Gene expression assays 3
  • Promoter assays 4
  • Further uses 5
  • See also 6
  • References 7
  • External links 8

Common reporter genes

To introduce a reporter gene into an organism, scientists place the reporter gene and the gene of interest in the same bacteria or prokaryotic cells in culture, this is usually in the form of a circular DNA molecule called a plasmid. It is important to use a reporter gene that is not natively expressed in the cell or organism under study, since the expression of the reporter is being used as a marker for successful uptake of the gene of interest.

Commonly used reporter genes that induce visually identifiable characteristics usually involve fluorescent and luminescent proteins. Examples include the gene that encodes jellyfish green fluorescent protein (GFP), which causes cells that express it to glow green under blue light, the enzyme luciferase, which catalyzes a reaction with luciferin to produce light, and the red fluorescent protein from the gene dsRed. The GUS gene has been commonly used in plants but luciferase and GFP are becoming more common.[1]

A common reporter in bacteria is the E. coli lacZ gene, which encodes the protein beta-galactosidase. This enzyme causes bacteria expressing the gene to appear blue when grown on a medium that contains the substrate analog X-gal. An example of a selectable-marker which is also a reporter in bacteria is the chloramphenicol acetyltransferase (CAT) gene, which confers resistance to the antibiotic chloramphenicol.

Gene name Gene product Assay Ref.
lacZ β-galactosidase enzyme assay or Histochemical
cat Chloramphenicol acetyltransferase Chloramphenicol acetylation
gfp Green fluorescent protein Fluorescent
rfp Red fluorescent protein microscopical, spectrophotometry [2]

Transformation and transfection assays

Many methods of promoter independent from that of the introduced gene of interest; the reporter gene can be expressed constitutively (that is, it is "always on") or inducibly with an external intervention such as the introduction of IPTG in the β-galactosidase system. As a result, the reporter gene's expression is independent of the gene of interest's expression, which is an advantage when the gene of interest is only expressed under certain specific conditions or in tissues that are difficult to access.

In the case of selectable-marker reporters such as CAT, the transfected population of bacteria can be grown on a substrate that contains chloramphenicol. Only those cells that have successfully taken up the construct containing the CAT gene will survive and multiply under these conditions.

Gene expression assays

Reporter genes can also be used to assay for the expression of the gene of interest, which may produce a protein that has little obvious or immediate effect on the cell culture or organism. In these cases the reporter is directly attached to the gene of interest to create a gene fusion. The two genes are under the same promoter elements and are transcribed into a single messenger RNA molecule. The mRNA is then translated into protein. In these cases it is important that both proteins be able to properly fold into their active conformations and interact with their substrates despite being fused. In building the DNA construct, a segment of DNA coding for a flexible polypeptide linker region is usually included so that the reporter and the gene product will only minimally interfere with one another.

Promoter assays

Reporter genes can be used to assay for the activity of a particular promoter in a cell or organism. In this case there is no separate "gene of interest"; the reporter gene is simply placed under the control of the target promoter and the reporter gene product's activity is quantitatively measured. The results are normally reported relative to the activity under a "consensus" promoter known to induce strong gene expression.

Further uses

A more complex usage of reporter genes on a large scale is in two-hybrid screening, which aims to identify proteins that natively interact with one another in vivo.

See also

References

  1. ^ Koo, J.; Kim, Y.; Kim, J.; Yeom, M.; Lee, I. C.; Nam, H. G. (2007). "A GUS/Luciferase Fusion Reporter for Plant Gene Trapping and for Assay of Promoter Activity with Luciferin-Dependent Control of the Reporter Protein Stability". Plant and Cell Physiology 48 (8): 1121–1131.  
  2. ^ Nordgren, I. K.; Tavassoli, A (2014). "A bidirectional fluorescent two-hybrid system for monitoring protein-protein interactions". Molecular BioSystems 10 (3): 485–90.  

External links

  • Research highlights and updated information on reporter genes.
  • Staining Whole Mouse Embryos for β-Galactosidase (lacZ) Activity
This article was sourced from Creative Commons Attribution-ShareAlike License; additional terms may apply. World Heritage Encyclopedia content is assembled from numerous content providers, Open Access Publishing, and in compliance with The Fair Access to Science and Technology Research Act (FASTR), Wikimedia Foundation, Inc., Public Library of Science, The Encyclopedia of Life, Open Book Publishers (OBP), PubMed, U.S. National Library of Medicine, National Center for Biotechnology Information, U.S. National Library of Medicine, National Institutes of Health (NIH), U.S. Department of Health & Human Services, and USA.gov, which sources content from all federal, state, local, tribal, and territorial government publication portals (.gov, .mil, .edu). Funding for USA.gov and content contributors is made possible from the U.S. Congress, E-Government Act of 2002.
 
Crowd sourced content that is contributed to World Heritage Encyclopedia is peer reviewed and edited by our editorial staff to ensure quality scholarly research articles.
 
By using this site, you agree to the Terms of Use and Privacy Policy. World Heritage Encyclopedia™ is a registered trademark of the World Public Library Association, a non-profit organization.
 


Copyright © World Library Foundation. All rights reserved. eBooks from Project Gutenberg are sponsored by the World Library Foundation,
a 501c(4) Member's Support Non-Profit Organization, and is NOT affiliated with any governmental agency or department.