Establishment of real-time TaqMan-fluorescence quantitative RT-PCR assay for detection and quantitation of ten kinds of porcine inflammation markers mRNA

Real-time PCR assays based on TaqMan probes for detection of porcine inflammation markers including interferon-α (IFN-α), interferon-β (IFN-β), retinoic acid-inducible gene 1 (RIG-1), toll-like receptor 9 (TLR-9), interferon regulatory factors (IRF-3, IRF-7), Janus kinase (JAK-1, JAK-2), signal transducers and activators of transcription (STAT-1, STAT-2) were established in this study. The results indicated that the established assays were highly specific and sensitive with a detection limit of 1.0 × 101 copies/μl, and coefficient of variations was less than three percent for both inter- and intra-assay. The established assays were used to detect mRNA levels of these inflammation markers and β-actin in swine testicle (ST) cells transfected with polyinosinic: polycytidylic acid (poly (I:C)). The results showed that the transcription levels (mRNA) of IFN-α, IFN-β, RIG-1 and IRF-7 were up-regulated in ST cells transfected with poly (I:C), and the transcription levels (mRNA) of TLR-9, IRF-3, JAK-1, JAK-2, STAT-1 and STAT-2 showed minimal change. The real-time PCR assays established in this study with high specificity, sensitivity and reproducibility could be used to quantify mRNA levels of porcine inflammation markers.