Quantitative PCR Analysis of Gene Expression

Total RNA extracted with RNeasy Mini Kit (Qiagen, Chatsworth, CA, USA) from the liver and kidneys was transcribed using ReverTra Ace qPCR RT Master Mix with gDNA Remover (FSQ30, Toyobo, Osaka, Japan). Quantitative PCR was performed using THUNDERBIRD Probe qPCR Mix (Toyobo) and StepOne plus (Thermo Fisher Scientific, Waltham, MA, USA) as described previously [4 (link),46 (link)], and the Taqman probes used are listed in Supplemental Table S1. The values obtained were used in the ΔΔCt method of relative quantification with normalization to 18S gene expression.

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Mishima E., Ichijo M., Kawabe T., Kikuchi K., Akiyama Y., Toyohara T., Suzuki T., Suzuki C., Asao A., Ishii N., Fukuda S, & Abe T. (2020). Germ-Free Conditions Modulate Host Purine Metabolism, Exacerbating Adenine-Induced Kidney Damage. Toxins, 12(9), 547.

Publication 2020

RNeasy Mini Kit ReverTra Ace qPCR RT Master Mix with gDNA Remover THUNDERBIRD Probe qPCR Mix StepOne plus

Gene expression Kidneys Liver

Corresponding Organization : Tohoku University

Other organizations : University of Tsukuba, Keio University

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Gene expression

control variables

Total RNA extraction method (RNeasy Mini Kit)
Reverse transcription method (ReverTra Ace qPCR RT Master Mix with gDNA Remover)
Quantitative PCR method (THUNDERBIRD Probe qPCR Mix, StepOne plus)
Normalization to 18S gene expression

controls

Positive control: Not mentioned
Negative control: Not mentioned

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