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首页> 外文期刊>Journal of Virological Methods >Establishment of real-time TaqMan-fluorescence quantitative RT-PCR assay for detection and quantitation of ten kinds of porcine inflammation markers mRNA
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Establishment of real-time TaqMan-fluorescence quantitative RT-PCR assay for detection and quantitation of ten kinds of porcine inflammation markers mRNA

机译：建立实时定量TaqMan荧光定量RT-PCR检测和定量分析十种猪炎症标志物mRNA的方法

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摘要

Real-time PCR assays based on TaqMan probes for detection of porcine inflammation markers including interferon-alpha (IFN-alpha), interferon-beta (IFN-beta), 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 x 10(1) copies/mu l, 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 beta-actin in swine testicle (ST) cells transfected with polyinosinic: polycytidylic acid (poly (I:C)). The results showed that the transcription levels (mRNA) of IFN-alpha, IFN-beta, RIG-1 and IRF-7 were up-regulated in ST cells transfected with poly (LC), 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
机译：基于TaqMan探针的实时PCR分析，用于检测猪炎症标志物，包括干扰素-α（IFN-alpha），干扰素-β（IFN-beta），视黄酸诱导基因1（RIG-1），toll​​样受体9（TLR-9），干扰素调节因子（IRF-3，IRF-7），Janus激酶（JAK-1，JAK-2），信号转导子和转录激活子（STAT-1，STAT-2）的建立这项研究。结果表明，已建立的测定具有高度特异性和敏感性，检出限为1.0 x 10（1）拷贝/微升，测定间和测定内变异系数均小于3％。建立的测定法用于检测用多肌苷酸：聚胞苷酸（poly（I：C））转染的猪睾丸（ST）细胞中这些炎症标志物和β-肌动蛋白的mRNA水平。结果表明，在转染了poly（LC）的ST细胞中，IFN-α，IFN-beta，RIG-1和IRF-7的转录水平（mRNA）上调，而TLR-如图9所示，IRF-3，JAK-1，JAK-2，STAT-1和STAT-2的变化很小。在这项研究中建立的具有高特异性，灵敏性和可重复性的实时PCR检测方法可用于定量猪炎症标志物的mRNA水平

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