Quantifying mRNA Expression by RT-qPCR

Total hepatic RNA was extracted with TRIzol^® (Invitrogen, Carlsbad, CA, USA) and reverse transcribed into cDNA with the High-Capacity cDNA Reverse Transcript Kit (Applied Biosystems) and diluted 1:10 with DNase and RNase-free H₂O. Real-time PCR was performed on an ABI 7900 using the SYBR Green assay (primer sequences were available on request) as described previously [4 (link)]. The delta-Ct method was used to calculate mRNA expression levels (expressed by ratio = 2^-deltaCt *100%), using Cyclophilin A as the control gene.

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Wu J., Cui W., Cai Q., Fei J., Zhang S.D., Han T.Q., Hu H, & Jiang Z.Y. (2016). The NPC1L1 Polymorphism 1679C>G Is Associated with Gallstone Disease in Chinese Patients. PLoS ONE, 11(1), e0147562.

Publication 2016

TRIzol High-Capacity cDNA Reverse Transcript Kit

Assay Cdna Cyclophilin a Dnase Gene control Mrna Primer Real time pcr Rnase Sybr green Trizol

Corresponding Organization : First Affiliated Hospital of Wannan Medical College

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Variable analysis

independent variables

Total hepatic RNA extraction method (TRIzol)

dependent variables

MRNA expression levels

control variables

Cyclophilin A as the control gene

controls

Positive control: Not specified
Negative control: Not specified

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