RNA was isolated from cell pellets using kit (catalogue no. 74104, Qiagen, USA), and then using a cDNA Reverse Transcription kit (Applied Biosystems, Thermo Fisher Scientific, Inc.), cDNA was synthesized. Quantitative PCR was performed using HotStart-IT® FideliTaq™ PCR Master Mix (2X) (catalog no. 71156, Affymetrix, USA). The effect of Aps on the expression of isolated genes families involved in the oxidative stress [Cat, SOD1 (Cu-Zn-SOD), and SOD2 (Mn-SOD)], inflammation [TNF-α and IL-6], apoptosis [Cas-3, Bax, and Bcl2] was investigated. The used primers are shown in Table 1 . PCR conditions were adjusted according to Huang et al.[32 (link)] CFX96 real-time system (Bio-Rad Laboratories, Inc.) was used for the reaction. Transcript levels were calculated and normalized of the expression of the targeted genes to the expression of GAPDH (internal reference gene). Experiments were conducted in triplicates.
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Quantitative Analysis of Oxidative Stress, Inflammation, and Apoptosis Genes
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Apoptosis
Bcl2
Cdna
Cell
Expression genes
Gapdh
Gene
Inflammation
Mn sod
Oxidative stress
Pellets
Primers
Reverse transcription
Sod2
Tnf α
Corresponding Organization :
Other organizations : King Abdulaziz University, Mansoura University, Northern Border University
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Variable analysis
dependent variables
- Expression of genes involved in oxidative stress (Cat, SOD1 (Cu-Zn-SOD), and SOD2 (Mn-SOD))
- Expression of genes involved in inflammation (TNF-α and IL-6)
- Expression of genes involved in apoptosis (Cas-3, Bax, and Bcl2)
- GAPDH (internal reference gene)
- No positive or negative controls were explicitly mentioned.
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