| Description | The RIG-I/NF-kB LUCPoter(TM) reporter cell line is designed to monitor the RIG-I-mediated NF-kB induction activity, and can be used for studying RIG-I signaling pathways as well as screening of agonists, antagonists or signaling inhibitors for RIG-I.
Contents: 3-4 x 10^6 cells Biosafety Level: 2 |
| Immunogen | The RIG-I/NF-kB LUCPorter(TM) reporter cell line is a stably transfected HEK 293T cell line which expresses human retinoic acid-induced protein-I (RIG-I) and an optimized Renilla luciferase reporter gene (RenSP) under the transcriptional control of an NF-kB response element. As a dsRNA helicase enzyme, RIG-I is encoded by the DDX58 gene. RIG-I is one of the RIG-I-like receptors (RLRs) that are a family of DExD/H box RNA helicases including RIG-I, MDA5 and LPG2, which play a major role in pathogen sensing of RNA virus infection to initiate and modulate antiviral immunity. RLR expression is typically maintained at low levels in resting cells but is greatly increased during inflammation, specifically with IFN exposure and after virus infection. RIG-I detects cytoplasmic dsRNA generated during viral replication unlike Toll-like receptor 3 (TLR3) which can detect phagocytosed dsRNA in endosomes. RIG-I also responds to poly(I:C), the synthetic analog of viral dsRNA. As shown in Figure 1, the RIG-I/NF-kB LUCPorter(TM) reporter cell line was specifically activated, in a dose response manner, by the poly(I:C) packed with Lipofectamine 2000, in which Lipofectamine mediated the cytoplasmic translocation of poly(I:C) (Figure 1). |
| Selection Agent | Blasticidin at 5 mg/ml and Puromycin at 3 mg/ml |
| Growth Properties | Adherent Morphology : Epithelial |
| RCL Type | Luciferase - (LUCPorter™) |
| Host | HEK293 |
| Dilutions |
|
| Application Notes | Refer to the following preparation protocol for the poly(I:C) ligand packed with Lipofectamine 2000: 1. A 500 ug (50 ul, 10 mg/ml) of poly(I:C) (NBP2-25288) was preincubated in 50 ul Opti-MEM (Life Technologies) for 5 min. Similarly, a 20 ul Lipofectamine 2000 (Life Technologies) was preincubated in 80 ul Opti-MEM for 5 min. After 5 min, they were combined together as a total volume of 200 ul and further incubated for 20 min at room temperature. 2. The poly(I:C) packed with Lipofectamine 2000 (A 200 ul total at 2.5 mg/ml) was then used to stimulate the RIG-I/NF-kB LUCPorter(TM) HEK 293T cell line as described in Figure 1A. |
| Readout System |
| Storage | Store in gas phase of liquid nitrogen. |
| Reconstitution Instructions | Complete Growth Medium: DMEM with 4.5 g/L glucose + 10% FBS + 4 mM L-glutamine + 1 mM sodium pyruvate + 100 units/ml penicillin + 0.1 mg/ml streptomycin (Note: The selection agents for this cell line are blasticidin at 5 ug/ml and puromycin at 3 ug/ml). |
