Total RNA was extracted using TRIzol Reagent (Invitrogen), and complement DNA (cDNA) was synthesized using a PrimeScript RT Reagent Kit (Takara Bio, San Jose, CA, USA). The RT-qPCR reactions were performed using the SYBR Premix Ex Taq II kit (Takara Bio) and detected on the ABI Prism 7500 HT sequence detection system (Applied Biosystems, Foster City, CA, USA). The primer information of matrix metallopeptidase (MMP) MMP2, MMP9, and β-actin can be found in the publication of Zhao et al. [2017] (Table S1 ) (23 (link)); that for MDV Eco Q fragment-encoded gene MEQ from Heidari et al. [2008] (26 (link)); that for BCL2 and BCL2L1 from Subramaniam et al. [2013] (27 (link)); that for TNFSF10 from Li et al. [2008] (28 (link)); and that for the DNMT3B gene from Tian et al. [2013] (Table S1 ) (29 (link)). The results were determined by the 2–ΔΔCt method (30 (link)).
Abi prism 7500 ht sequence detection
Manufactured by Thermo Fisher Scientific
Sourced in United States, Germany
The ABI Prism 7500 HT sequence detection system is a real-time PCR instrument designed for high-throughput analysis. It can perform quantitative, qualitative, and genotyping analysis of DNA and RNA samples.
Automatically generated - may contain errors
Lab products found in correlation
Sybr premix ex taq 2 kit, by Takara Bio (1 mentions)
Trizol reagent, by Thermo Fisher Scientific (1 mentions)
Primescript rt reagent kit, by Takara Bio (1 mentions)
Sybr green pcr master mix, by Qiagen (1 mentions)
Reverse transcription kit, by Qiagen (1 mentions)
Trizol, by Thermo Fisher Scientific (1 mentions)
Dnase, by Thermo Fisher Scientific (1 mentions)
2 protocols using abi prism 7500 ht sequence detection
1
RT-qPCR Analysis of Gene Expression
2
Quantitative RT-PCR Analysis of Gut Gene Expression
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Total RNA was extracted from jejunal tissues using Trizol (Invitrogen, USA). RNA samples were processed with DNase (Applied Biosystems, Darmstadt, Germany) for at least 1 hour before being transformed into cDNA according to the manufacturer’s instructions using a reverse transcription kit (Qiagen, Hilden, Germany). qRT-PCR was carried out using an ABI Prism 7500HT sequence detection system (Applied Biosystems, Darmstadt, Germany) with Qiagen’s SYBR green PCR master mix (Hilden, Germany). The mRNA genes for interferon- γ (IFN- γ), inducible nitric oxide synthase (iNOs), interleukin-10 (IL-10), NF-kB, as well as beta-actin (β-actin) as a housekeeping control were studied using SYBR green (Hilden, Germany). All primer assays for qRT-PCR were obtained from Qiagen (Hildan, Germany). Real-time qPCR amplification and analysis were carried out using the Bio-Rad IMark Microplate Reader SW 1.04.02.E. The Ct method (2−ΔΔCT) described by Livak and Schmittgen (35 (link)) was used to analyze differences in gene expression.
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