Quantitative RT-PCR for Gene Expression

One microgram of total RNA was reverse transcribed to cDNA [37 (link)] and amplified with gene-specific primers labeled with Power SYBR Green master mix (Applied Biosystems, Foster City, CA, USA) using an QuantStudio 3 Real time PCR (Applied Biosystems). Samples were normalized to the expression of 18S rRNA using the comparative CT method [82 (link)]. Sequences for the primers (Il1b, Tlr4, Nalp3) can be found in Table S1 or in previously published works (Kc, Mcp1, Cox2, Il6, 18S) [36 (link),37 (link),40 (link)].

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Romo D., Velmurugan K., Upham B.L., Dwyer-Nield L.D, & Bauer A.K. (2019). Dysregulation of Gap Junction Function and Cytokine Production in Response to Non-Genotoxic Polycyclic Aromatic Hydrocarbons in an In Vitro Lung Cell Model. Cancers, 11(4), 572.

Publication 2019

Power SYBR Green master mix QuantStudio 3 Real time PCR

18s rrna Cdna Cox2 Gene Il1b Mcp1 Primers Real time pcr Sybr green

Corresponding Organization : University of Colorado Anschutz Medical Campus

Other organizations : Michigan State University

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Variable analysis

independent variables

Total RNA concentration (1 microgram)

dependent variables

MRNA expression levels of Il1b, Tlr4, Nalp3, Kc, Mcp1, Cox2, Il6

control variables

18S rRNA expression (used for normalization)

controls

Positive control: Not explicitly mentioned.
Negative control: Not explicitly mentioned.

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