16.11 : Genes Repórteres

Reporter genes are a type ofprotein-coding genethat are often tagged to a gene of interest. Once inside a target cell, reporter genes usually produce visually identifiable characteristics like fluorescence and luminescence when expressed along with the gene of interest. Thus, reporter genes “report” the presence or absence of genes of interest in an organism, determine the gene expression pattern, or track the physical location of a DNA segment or protein in the cell.

Commonly used reporter genes are - GFP (green fluorescent protein gene), lacZ (β- galactosidase gene), RFP (red fluorescent protein gene), and Luc (luciferase gene). The cells expressing GFP glow green when excited by wavelengths of blue to the ultraviolet range, while those expressing RFP glow red when excited by wavelengths of 488 nm or 532 nm. The cells expressing the Luc gene produce luciferase enzymes that catalyze a reaction with luciferin to produce light. The lacZ gene is the most common reporter gene used in E.coli. It produces an enzyme β-galactosidase that causes the bacteria to appear blue when grown in a media containing X-gal substrate.

Reporter genes to study promoter activity

Besides the use of reporter genes in studying gene expression and localization, they can also help in studying the strength of promoters. This is done by placing a reporter gene downstream of the promoter in a plasmid and then introducing it into the mammalian cell. The expression of the reporter gene monitors the activity of the promoter. If the reporter gene expression is high, the promoter is said to be a strong promoter; it’s weak if the reporter gene expression is low.

Reporter gene, when placed downstream of the promoter in a plasmid, also helps in measuring transfection efficiency. The recombinant plasmid is transfected into a mammalian cell, and its expression shows that the cell is successfully transfected. The amount of protein produced by the recombinant plasmid shows its transfection efficiency.