Reporter Gene Expression for Monitoring Microorganisms in the Environment

互联网2014-02-13

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Reporter gene is a generic term that is used to describe genes whose expression is indicative of the presence of a particular cell, a specific event occurring within that cell, or a reflection of the expression of other, usually targeted genes of interest. For example, the presence of such a gene may make an organism standout to help in identification, as will be described in this chapter, or it could be that a gene is only expressed under certain conditions, e.g., during different developmental stages (1 ), environmental stresses (2 ,3 ) , or DNA repair as a result of induction of the SOS response to DNA damage (4 ). Naturally-occurring reporter genes can be used to avoid the need for genetic engineering by selecting a unique property of an organism and using it as the reporter. However, by far the commonest approach is to introduce a reporter gene into a cell to allow detection of a particular organism or to monitor its activity. The reporter gene can be located on a plasmid or to increase the stability of the gene within the cell it can be incorporated into the bacterial chromosome. There are a wide variety of reporter genes available, and all have particular properties that make them more or less useful depending on the application (Table 1 ). The choice of a particular reporter gene is made on the basis of the environment into which the organism is to be released, how the gene is to be detected, and what factors might interfere with the detection of the gene and/or its protein product.

Table 1 Examples of Different Types of Reporter Genes

Method of detection	Source	Reporter gene	Gene product	Mode of detection	Reference
Resistance	Various plasmids	Antibiotic resistance genes, e.g., catP amp	For example, chloramphenicol acetyltransferase, β-lactamase	Selective culture in presence of antibiotic, herbicide	32
	Pseudomonas putida EEZ15	Herbicide resistance	Unknown	or metal	6
	Various plasmids	Heavy metal resistance, e.g., merA	For example, mercuric reductase		5
Fluorescence	Fluorescent marine bacteria, e.g., Vibrio fischerii, V. harveyi, Photobacterium spp.	lux	Luciferase	Scintillation counter, luminometry, CCD (chargecoupled device) camera,	*3,4,9,19–26*
	Fire Fly	luc		X-ray film,	33
	Click beetle	rluc		microscopy	1
	Aequorea victoria (jellyfish) Renilla reniformis (sea pansy)	gfP	Green fluorescent protein (GFP)	fluorescent microscopy, flow cytometry,	*15–18*
	V. fischerii	yfp	Yellow fluorescent protein (YFP)	confocal microscopy	38
Chromogenic	E. coli, Bacillus stearothermophilus	lacZY, BgaB	β-Galactosidase	Enzyme assay in presence of	*2,7,8,9 35*
	E. coli	gusA, uidA	β-Glucuronidase	appropriate substrate, e.g., X-Gal, catachol	31
	TOL plasmid pWWO of Pseudomonas putida	xylE	Catechol 2,3-dioxygenase	spectrophotometry, visually	*10–14*

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