How does the luciferase assay work?
A Luciferase Reporter Assay. When this protein activates transcription, the cell will produce luciferase enzyme. After the addition of a lysis buffer and a substrate, a luminometer quantifies the luciferase activity. If your protein activates the expression of the target gene, the amount of signal produced increases.
What is Renilla luciferase assay?
Reliable, Renilla Luciferase Assay for Primary Expression or Reporter Normalization. Renilla Luciferase Assay System is designed to provide a fast, sensitive method of detecting luciferase from sea pansy (Renilla reniformis).
What does a dual luciferase assay do?
The DLR assay exploits the differing biochemical requirements for luminescence of the firefly (Photinus pyralis) and sea pansy (Renilla reniformis) luciferase proteins (31). This allows for the sequential quantitative measurement of both luciferase activities in a single protein extract.
How do you use luciferase reporter assay?
To perform the reporter assay, you clone the regulatory region of your gene-of-interest (X) upstream of the luciferase gene in one of these expression vectors, introduce that resulting vector DNA into cells, and let the cells grow for a period of time to allow transcription and translation to occur.
What is the difference between Renilla and firefly luciferase?
What is the Difference Between Firefly and Renilla Luciferase? Firefly luciferase assays uses luciferin in the presence of oxygen, ATP and magnesium to produce light (Green/Yellow, 550-70 nM), while Renilla luciferase assays (SCT153) requires only coelenterazine and oxygen to produce light (Blue, 480 nM).
What color is Renilla luciferase?
Renilla luciferase enzyme/protein (RL), purified from sea pansy (Renilla reniformis), is a bioluminescent soft coral that displays blue-green bioluminescence upon mechanical stimulation.
What is luciferase RNA?
Luciferase Control RNA is an uncapped in vitro-transcribed RNA containing a 30-base poly(A) tail that produces functional luciferase when translated. Control reactions are monitored easily by a luciferase assay for the production of luminescence generated from the full-length luciferase.
What is dual luciferase reporter system?
The Dual-Luciferase® Reporter (DLR™) Assay System provides an efficient means of performing two reporter assays. In the DLR™ Assay, the activities of firefly (Photinus pyralis) and Renilla (Renilla reniformis or sea pansy) luciferases are measured sequentially from a single sample.
What is Nano luciferase?
NanoLuc luciferase (NLuc) is the newest commercially available luciferase enzyme. NLuc is a 19.1 kDa luciferase enzyme that relies on the substrate furimazine to produce high intensity, glow-type luminescence. 4. Recently, NLuc was shown to exhibit several properties superior to traditional luciferases.
What is the luciferase reaction?
Luciferase is an enzyme. An enzyme is a chemical (called a catalyst) that interacts with a substrate to affect the rate of a chemical reaction. The interaction of the luciferase with oxidized (oxygen-added) luciferin creates a byproduct, called oxyluciferin. More importantly, the chemical reaction creates light.
Why is luciferase used as a reporter gene?
The luciferase reporter assay is commonly used as a tool to study gene expression at the transcriptional level. It is widely used because it is convenient, relatively inexpensive, and gives quantitative measurements instantaneously.
What is luciferase reporter gene?
A commonly used reporter gene is the luciferase gene from the firefly Photinus pyralis. This gene encodes a 61-kDa enzyme that oxidizes D-luciferin in the presence of ATP, oxygen, and Mg(++), yielding a fluorescent product that can be quantified by measuring the released light.
How do you write a reporter gene?
To introduce a reporter gene into an organism, scientists place the reporter gene and the gene of interest in the same DNA construct to be inserted into the cell or organism. For bacteria or prokaryotic cells in culture, this is usually in the form of a circular DNA molecule called a plasmid.
What is the role of luciferase?
Luciferases are proteins with enzymatic activity that, in the presence of ATP, oxygen, and the appropriate substrate (typically luciferin), catalyze the oxidation of the substrate in a reaction that results in the emission of a photon.
Why is luciferase important?
The power of luciferase has been harnessed by scientists to devise reactions whose light output is used to monitor biological processes including gene expression, biomolecular binding, and cell viability.
Who named luciferase?
Raphaël Dubois
Luciferase is a generic term for the class of oxidative enzymes that produce bioluminescence, and is usually distinguished from a photoprotein. The name was first used by Raphaël Dubois who invented the words luciferin and luciferase, for the substrate and enzyme, respectively.
What is dual luciferase reporter assay?
How do reporter gene assays work?
Reporter gene assays are typically used to measure the regulatory ability of an unknown DNA-sequence. This is done by linking the unknown promoter sequence to an easily detectable reporter gene whose product can be easily detected and quantifiably measured.