Total cellular mRNA was extracted using an RNeasy Mini Kit (QIAGEN Sciences, Germany) according to the manufacturer’s protocol. cDNA was synthesized using a PrimeScriptTM II 1st Strand cDNA Synthesis Kit (Takara, China). Quantitative real-time PCR (qPCR) was performed using Power SYBR Green PCR Master Mix reagents (Takara, China). All data were acquired using a LightCycler 480 real-time PCR system (Roche, USA). The sequences of the RT-qPCR primers for amplification of TGEV, CXCL9, CXCL10, poIRF1, IFN-γ, and GAPDH transcripts are listed in Table 1 .
Power sybr green pcr master mix reagents
Manufactured by Takara Bio
Sourced in China, United States
Power SYBR Green PCR Master Mix is a ready-to-use solution for performing real-time PCR amplification using the SYBR Green I dye. It contains all necessary components for efficient and sensitive real-time PCR, including DNA polymerase, dNTPs, and SYBR Green I fluorescent dye.
Automatically generated - may contain errors
2 protocols using power sybr green pcr master mix reagents
1
Quantitative RT-qPCR Analysis of Immune Genes
2
Quantitative PCR for PEDV and miRNA
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Total cellular RNA was isolated using an RNeasy Mini kit (Qiagen Sciences, Hilden, Germany) according to the manufacturer's instructions. Total RNA was extracted and reverse transcribed as previously described (Ma et al., 2018 (link)). For miRNA reverse transcription, cDNA was prepared with a miRNA First Strand cDNA Synthesis kit (Sangon Biotech, Shanghai, China). qPCR was conducted in triplicate with Power SYBR Green PCR Master Mix reagents (Takara) on a LightCycler480 II system (Thermo Fisher Scientific, Waltham, MA, USA) as previously described (Ma et al., 2018 (link)). The miRNA expression levels were normalized to the internal control of U6. The sequences of RT-qPCR primers for PEDV, IFN-λ, SOCS1, IFIT1, ISG15, MxA, GAPDH, miR-30c, and Uni-miR transcription are listed in Table 2 . The results are presented as the means ± SEM from three separate trials.
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