5'ppp-dsRNA/LyoVec™

RIG-I Agonist - complexed with transfection reagent

5’ triphosphate double stranded RNA (5’ ppp-dsRNA) is a synthetic ligand for retinoic acid-inducible protein I (RIG-I).

RIG-I is a cytosolic pattern recognition receptor that senses pathogen-associated molecular patterns (PAMPs) on viral RNA and triggers an antiviral immune response by the activation of type-I interferons (IFN-α and -β) [1].

RIG-I shares the identification of the dsRNA structure with another sensor MDA-5 (recognition of poly(I:C)/LyoVec™) and is specifically activated by the uncapped 5’ triphosphate moiety on viral RNA [2].

5’ ppp-dsRNA is complexed with the cationic lipid LyoVec™ to facilitate its uptake.

1. Yoneyama M. & Fujita T., 2007. Function of RIG-I-like Receptors in Antiviral Innate Immunity. J. Biol. Chem. 282: 15315-8.
2. Hornung V. et al., 2006. 5'-Triphosphate RNA is the ligand for RIG-I. Science. 314(5801):994-7.

Specifications

Specificity: RIG-I agonist

Working Concentration: 300 ng- 1 μg/ml

Endotoxin level:  EndoFit™ (<0.001 EU/µg)

Solubility: 0.05 mg/ml in water

Formulation: complexed with the transfection reagent LyoVec™

5’ppp-dsrna
5’- pppGCAUGCGACCUCUGUUUGA -3’ (19 mer)
3’- CGUACGCUGGAGACAAACU -5’ (19 mer)

Contents

5’ppp-dsRNA/LyoVec™ is provided lyophilized.

This product is available in two sizes:

• tlrl-3prnalv :

- 25 μg 5’ppp-dsRNA/LyoVec™.
- 2 ml sterile apyrogenic water.

• tlrl-3prnalv-100:

- 100 μg (4x25μg) 5’ppp-dsRNA/LyoVec™.
- 10 ml sterile apyrogenic water.

Note: Each vial contains 25 μg of 5’ppp-dsRNA complexed with 50 μg LyoVec™.

5’ppp-dsRNA/LyoVec™ is shipped at room temperature.

Store lyophilized 5’ppp-dsRNA/Lyovec at -20°C.

Lyophilized product is stable for 1 year when stored -20°C.

Description

Induction of RIG-I with 5’ppp-dsRNA/LyoVec™ can be studied in a variety of cells expressing RIG-I.

If your cell line does not naturally express the RIG-I gene, transfect with a plasmid expressing a RIG-I gene, such as pUNO-hRIG-I or pUNO-mRIG-I.

RIG-I activation can be easily monitored using B16-Blue™ ISG cells, an IFN Regulatory Factor (IRF)-Inducible SEAP reporter cell line. In these cells, activation of RIG-I triggers the secretion of IFNs that results in the production of SEAP.

A protocol for the induction of RIG-I using B16-Blue™ ISG cells is given below:

• - Resuspend 5’ppp-dsRNA/LyoVec™ in water.
• - Stimulate cells with 1 - 10 µg/ml 5’ppp-dsRNA/LyoVec™ for 16 - 24 hours.

Note: The use of 5’ppp-dsRNA Control (as a negative control) and Poly(I:C)/LyoVec™ (as a positive control) is highly recommended.

• - Monitor induction of RIG-I by measuring the levels of SEAP in the cell culture supernatant using QUANTI-Blue™, a SEAP detection reagent.