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RNA isolation, reverse transcription and qPCR

AZ Alzbeta Zinkova
IB Iva Brynychova
AS Alexander Svacina
MJ Marie Jirkovska
MK Marie Korabecna
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RNA was isolated by GenEluteTM Mammalian Total RNA Miniprep Kit (Sigma Aldrich, USA). Extracted RNA was reverse-transcribed into cDNA by HyperScriptTM One-step RT-PCR Master mix (GeneAll, Korea). QPCR was performed in LightCycler®480 System (Roche Applied, USA) with Assays-on-Demand TaqMan® Gene Expression Assay Mix (Life Technologies-Applied Biosystems,USA) for PGK1 Hs99999906_m1 (Phosphoglycerate Kinase-1) as a reference gene and TNF-α Hs01113624_g1. The expressions of both genes were examined in all samples, the levels of TNF-α mRNA expression were given as = 2−∆Ct related to the reference gene PGK1.

Copyright and License information: The Author(s) ©2017
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