Total RNA was extracted using Trizol reagent (Thermo, USA). Quantitative analysis of gene expression was performed using the HiScript 1st Strand cDNA Synthesis Kit and SYBR-Green I Real-Time PCR kit (Vazyme, China) on an ABI 7500 real-time PCR Detection System (Thermo). The expression levels were normalized to β-actin as a reference gene. The primer sequences used for mRNA analysis are provided in Supplementary Table 1 .
Sybr green 1 real time pcr kit
Manufactured by Vazyme
Sourced in China
The SYBR-Green I Real-Time PCR kit is a laboratory equipment product designed for real-time PCR applications. It utilizes the SYBR-Green I dye to detect and quantify DNA amplification in real-time.
Automatically generated - may contain errors
Lab products found in correlation
Trizol reagent, by Thermo Fisher Scientific (4 mentions)
Hiscript 1st strand cdna synthesis kit, by Vazyme (3 mentions)
Miscript sybr green pcr kit, by Qiagen (2 mentions)
Mirna primers, by Qiagen (2 mentions)
Miscript 2 rt kit, by Qiagen (2 mentions)
Abi 7500 real time pcr detection system, by Thermo Fisher Scientific (1 mentions)
Abi 7500, by Thermo Fisher Scientific (1 mentions)
Mirneasy serum plasma kit, by Qiagen (1 mentions)
Cfx96 touch real time pcr detection system, by Bio-Rad (1 mentions)
Revertaid first strand cdna synthesis kit, by Thermo Fisher Scientific (1 mentions)
Fluorescence thermal cycler, by Bio-Rad (1 mentions)
4 protocols using sybr green 1 real time pcr kit
1
Quantitative Gene Expression Analysis
2
Quantitative Analysis of miRNA and mRNA
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Total RNA was isolated from cells by using Trizol reagent (Thermo Fisher, USA). Total RNA in serum was purified by using an miRNeasy serum/plasma kit (QIAGEN, Germany) according to the manufacturer’s procedures. Quantitative analyses of miRNA were performed with a miScript II RT kit and miScript SYBR green PCR kit (QIAGEN, Germany). Quantitative analyses of mRNAs were conducted with HiScript 1st strand cDNA synthesis kit and SYBR-green I real-time PCR kit (Vazyme, China) on a real-time PCR detection system (ABI 7500, Thermo Fisher). The relative expression levels of miRNAs and mRNAs were normalized to the expression of RNU6B and β-actin, respectively. miRNA primers were supplied by QIAGEN. All the primer sequences for mRNAs are listed in Table S2 .
3
Quantitative Analysis of miRNAs and mRNAs in MSCs
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Total RNA of MSCs were extracted with Trizol Reagent (Invitrogen, USA). miRNA qRT-PCR were performed with miScript II RT Kit and miScript SYBR Green PCR Kit (Qiagen, Germany). qRT-PCR of cytokine mRNAs were conducted with HiScript 1st Strand cDNA Synthesis Kit and SYBR-Green I Real-Time PCR kit (Vazyme Biotech Co., Ltd, China). The amplification fluorescence signals were detected by CFX96 Touch™ Real-Time PCR Detection System (Bio-Rad, USA). The relative expression levels of miRNAs and mRNAs were normalized to the expression of RNU6B and β-actin, respectively. miRNA primers were supplied by Qiagen. All the primers sequences and qRT-PCR conditions for mRNAs are listed in Supplementary Table 1 .
4
Quantitative RT-PCR Analysis of Gene Expression
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Total RNA was extracted from the cells using TRIzol® reagent (Life Technologies) according to the manufacturer’s instructions, and an equal amount of RNA was reverse transcribed using the RevertAid First Strand cDNA Synthesis kit (Fermentas, Glen Burnie, MD, USA). RT-qPCR was performed to detect the changes in mRNA expression using the SYBR-Green I Real-Time PCR kit (Vazyme Biotech Co., Ltd., Nanjing, China) and the Bio-Rad fluorescence thermal cycler (Bio-Rad Laboratories, Hercules, CA, USA). The relative mRNA expression was normalized to the insert control gene, β-actin, according to the manufacturer’s instructions. The primers used in the present study were produced by Invitrogen (Life Technologies). All primer sequences and RT-qPCR conditions are listed in Table I .
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