Reporter gene
A reporter gene is a gene that can be used to track the expression of other genes. The reporter gene is usually introduced under the same promoter after the gene to be examined . In some cases, the activity of the promoter itself is of interest, without a target gene. A reporter enzyme , a fluorescent reporter protein or a detectable antigen is sometimes also expressed as a fusion protein. Fluorescent reporter genes are particularly important in fluorescence microscopy .
Well-known reporter systems
- the cat gene, coding for a chloramphenicol acetyltransferase (CAT)
- the gfp gene from Aequorea victoria , codes for a green fluorescent protein (GFP)
- the gusA gene from Escherichia coli , codes for a β-glucuronidase (GUS) (formerly known as uidA gene)
- the lacZ gene from Escherichia coli , codes for a β-galactosidase (β-Gal), mostly for blue-white selection
- the luciferase genes from Photinus pyralis and Renilla reniformis , code for bioluminescence enzymes
- the luciferase gene from Oplophorus gracilirostris , a deep-sea shrimp , codes for a bioluminescent enzyme; NanoLuc is the genetically engineered 19 kD subunit of Oploporus luciferase
- the Gaussia luciferase gene from Gaussia princeps , a copepod , codes for a bioluminescent enzyme
- the aequorin gene from bioluminescent jellyfish of the genus Aequorea , codes for a photoprotein that is used as a calcium sensor
- the phoA gene from Escherichia coli , codes for an alkaline phosphatase (AP)
The cDNAs for these proteins are cloned into suitable expression vectors in the corresponding reporter systems .
Example from plant physiology
The cDNA sequence of the β-glucuronidase (GUS) enzyme is added after the promoter sequence of the gene to be examined, and then transformed into a plant. Depending on the type of promoter, glucuronidase is then expressed in different cell types, development stages or environmental conditions, which produces a blue dye after the addition of the artificial colorless substrate X-Gluc . With this system one can track gene expression on organ, tissue and cell level.
literature
- Kain, SR & Ganguly S. (2001): Overview of Genetic Reporter Systems In: Current Protocols in Molecular Biology 2001 May; Chapter 9: Unit 9.6 PMID 18265284
- Shuman, HA & Silhavy, TJ (2003): The art and design of genetic screens: Escherichia coli. In: Nat. Rev. Genet. Vol. 4, pp. 419-31. PMID 12776212
See also
Individual evidence
- ↑ Bat-Erdene Jugder, Jeffrey Welch, Nady Braidy, Christopher P. Marquis: Construction and use of aCupriavidus necatorH16 soluble hydrogenase promoter (PSH) fusion togfp (green fluorescent protein) . In: PeerJ . tape 4 , July 26, 2016, ISSN 2167-8359 , doi : 10.7717 / peerj.2269 ( peerj.com [accessed November 1, 2017]).
- ↑ MP Hall, J. Unch, BF Binkowski, MP Valley, BL Butler, MG Wood, P. Otto, K. Zimmerman, G. Vidugiris, T. Machleidt, MB Robers, HA Benink, CT Eggers, MR Slater, PL Meisenheimer , DH Klaubert, F. Fan, LP Encell, KV Wood: Engineered luciferase reporter from a deep sea shrimp utilizing a novel imidazopyrazinone substrate. In: ACS chemical biology. Volume 7, number 11, November 2012, pp. 1848-1857, doi : 10.1021 / cb3002478 , PMID 22894855 , PMC 3501149 (free full text).
- ↑ BA Tannous, DE Kim, JL Fernandez, R. Weissleder, XO Breakefield: Codon-optimized Gaussia luciferase cDNA for mammalian gene expression in culture and in vivo. In: Molecular therapy: the journal of the American Society of Gene Therapy. Volume 11, Number 3, March 2005, pp. 435-443, doi : 10.1016 / j.ymthe.2004.10.016 , PMID 15727940 .
- ↑ H. Tanahashi, T. Ito, S. Inouye, FI Tsuji, Y. Sakaki: Photoprotein aequorin: use as a reporter enzyme in studying gene expression in mammalian cells. In: Genes. Volume 96, Number 2, December 1990, pp. 249-255, PMID 2269434 .