Quantitative Real-Time PCR Analysis of Liver Gene Expression

Total RNA was extracted from liver tissues using Trizol Reagent (Invitrogen, Carlsbad, CA). M-MLV Transcriptase was used for reverse transcription according to the manufacturer’s instructions (Thermo, Waltham, USA). The qRT-PCR was run in a thermocycler (iQ5 Bio-Rad, Hercules, CA) with the SYBR Green PCR premix (TaKaRa, Dalian, China) as described previously^{17 (link)}. The PCR primers used herein are noted in Table 1. The results were calculated by the 2^−△△Ct method.Table 1

Sequences of the qRT-PCR primers.

Gene	Genbank accession	Forward primer	Reverse primer
Acta2	NM_007392.3	AAGAGCATCCGACACTGCTGAC	AGCACAGCCTGAATAGCCACATAC
COL1A1	NM_007742.4	CAGGGTATTGCTGGACAACGTG	GGACCTTGTTTGCCAGGTTCA
IFN-γ	NM_008337.4	TAGCCAAGACTGTGATTGCGG	AGACATCTCCTCCCATCAGCAG
TNF-α	NM_013963.3	AAGCCTGTAGCCCACGTCGTA	AGGTACAACCCATCGGCTGG
IL-6	NM_031168.2	CCACTTCACAAGTCGGAGGCTTA	CCAGTTTGGTAGCATCCATCATTTC
IL-4	NM_021283.2	ACAGGAGAAGGGACGCCAT	GAAGCCCTACAGACGAGCTCA
IL-5	NM_010558.1	TGGGGGTACTGTGGAAATGC	CCACACTTCTCTTTTTGGCGG
IL-10	NM_010548.2	GCCAGAGCCACATGCTCCTA	GATAAGGCTTGGCAACCCAAGTAA
IL-13	NM_008355.3	CCCTGGATTCCCTGACCAAC	GGAGGCTGGAGACCGTAGT
IL-17A	NM_010552.3	ACCGCAATGAAGACCCTGAT	TCCCTCCGCATTGACACA
TGF-β1	NM_011577.2	GTGTGGAGCAACATGTGGAACTCTA	CGCTGAATCGAAAGCCCTGTA
FGL2	NM_008013.4	TGGACAACAAAGTGGCAAATCT	TGGAACACTTGCCATCCAAA
GAPDH	NM_001289726.1	CATGGCCTTCCGTGTTCCTA	CCTGCTTCACCACCTTCTTGAT

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Zhang C., Shao Y., Yang S., Bi X., Li L., Wang H., Yang N., Li Z., Sun C., Li L., Lü G., Aji T., Vuitton D.A., Lin R, & Wen H. (2017). T-cell tolerance and exhaustion in the clearance of Echinococcus multilocularis: role of inoculum size in a quantitative hepatic experimental model. Scientific Reports, 7, 11153.

Publication 2017

Trizol Reagent M-MLV Transcriptase SYBR Green PCR premix

Primers Reverse transcription Sybr green Transcriptase Trizol

Corresponding Organization :

Other organizations : First Affiliated Hospital of Xinjiang Medical University, Xinjiang Medical University, University of Science and Technology of China, Methodology and Quality of Life Unit in Oncology

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Expression levels of Acta2, COL1A1, IFN-γ, TNF-α, IL-6, IL-4, IL-5, IL-10, IL-13, IL-17A, TGF-β1, and FGL2 genes

control variables

None explicitly mentioned

controls

No positive or negative controls were explicitly mentioned in the protocol.

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