RNA Extraction, cDNA Synthesis, and qPCR Analysis

Total RNA was extract by using the TripleXtractor reagent (Bio-Cell) as indicated by the supplier, and the AMV Reverse Transcriptase kit (Promega) was used to produce cDNA following the manufacturer’s recommendations, by using 2 μg of total RNA. Quantitative PCR reactions were performed by using the Bio-Rad CFX96 qPCR thermocycler. The primer pair sequences for selected amplicons were designed using the online IDT PrimerQuest Tool software (IDT; https://eu.idtdna.com/Primerquest/Home/Index). Primer sequences are available in Supplementary Table 1. Gapdh mRNA level was used as an internal control and results were expressed as previously described^{56 (link)}.

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Dematteis G., Vydmantaitė G., Ruffinatti F.A., Chahin M., Farruggio S., Barberis E., Ferrari E., Marengo E., Distasi C., Morkūnienė R., Genazzani A.A., Grilli M., Grossini E., Corazzari M., Manfredi M., Lim D., Jekabsone A, & Tapella L. (2020). Proteomic analysis links alterations of bioenergetics, mitochondria-ER interactions and proteostasis in hippocampal astrocytes from 3xTg-AD mice. Cell Death & Disease, 11(8), 645.

Publication 2020

AMV Reverse Transcriptase kit CFX96 qPCR thermocycler

Cdna Cell Gapdh Mrna Primer Promega Reverse transcriptase

Corresponding Organization :

Other organizations : Università degli Studi del Piemonte Orientale “Amedeo Avogadro”, Lithuanian University of Health Sciences

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Variable analysis

independent variables

Total RNA extraction using TripleXtractor reagent
CDNA production using AMV Reverse Transcriptase kit

dependent variables

Quantitative PCR expression levels of selected amplicons

control variables

Gapdh mRNA level as an internal control

controls

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

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