pNiFty3-AN-Rluc

InvivoGen has designed pNiFty3, a collection of inducible reporter plasmids, to monitor pattern recognition receptor (PRR) activation and cytokine signaling upon ligand stimulation.

The pNiFty3- AN-Rluc plasmid features an AP-1/NF-κB-inducible Renilla luciferase (Rluc) reporter gene under the control of an engineered murine interferon beta (mIFN-β) promoter. This promoter comprises five AP-1 and five NF-κB repeated transcription factor binding sites (TFBS) to enhance the AP-1/NF-κB-mediated transcription. Of note, the Renilla luciferase remains intracellular and requires cell lysis in order to measure bioluminescence. The subsequent expression of Rluc upon receptor activation is readily measurable after cell lysis when using QUANTI-Luc™ 4 Renilla, a Renilla luciferase detection kit, that also includes a lysis buffer. This plasmids are selectable with Zeocin® in both E. coli and mammalian cells and can be used to generate stable clones.

Specifications

Transcription factor binding sites: AP-1 (5x) NF-κB (5x)
Minimal Promoter: mouse IFNβ promoter
Selection: Zeocin®
Reporter Gene: Renilla luciferase

These products are covered by a Limited Use License (See Terms and Conditions).

Contents

• 20 µg of lyophilized DNA
• 1 ml of Zeocin® (100 mg/ml)

Product is shipped at room temperature.

Details

Plasmid features:

AP-1-5x NF-κB-5x mIFN-β is an engineered murine interferon beta (mIFN-β) promoter comprising different positive regulatory domains that bind transcription factors such as NF-kB, IRF3 and IRF7 [1]. This minimal promoter is specific to AP-1 and NF-κB due to the addition of five AP-1 repeated transcription factor binding sites (TFBS) and five NF-κB TFBS, respectively. This feature enhances the AP-1/ NF-κBmediated transcription of the Rluc reporter gene.

The Renilla luciferase (Rluc) gene encodes for an intracellular luciferase from the sea pansy renilla reniformis. This enzyme catalyzes coelenterazine oxidation leading to bioluminescence and the production of light emission peaking at 480 nm [2]. After cell lysis, the activity can be evaluated using QUANTI-Luc™ 4 Renilla, an assay reagent containing all the components required to quantitively measure the activity of Renilla luciferase and other coelenterazine-utilizing luciferases.