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Selection of reference genes for normalization of real-time PCR data in minipig heart failure model and evaluation of TNF-α mRNA expression

Paper ID Volume ID Publish Year Pages File Format Full-Text
23824 43476 2011 8 PDF Available
Title
Selection of reference genes for normalization of real-time PCR data in minipig heart failure model and evaluation of TNF-α mRNA expression
Abstract

Real-time PCR is the benchmark method for measuring mRNA expression levels, but the accuracy and reproducibility of its data greatly depend on appropriate normalization strategies. Though the minipig model is largely used to study cardiovascular disease, no specific reference genes have been identified in porcine myocardium. The aim of the study was to identify and validate reference gene to be used in RT-PCR studies of failing (HF) and non-failing pig hearts. Eight candidate reference genes (GAPDH, ACTB, B2M, TBP, HPRT-1, PPIA, TOP2B, YWHAZ) were selected to compare cardiac tissue of normal (n = 4) and HF (n = 5) minipigs. The most stable genes resulted: HPRT-1, TBP, PPIA (right and left atrium); PPIA, GAPDH, ACTB (right ventricle); HPRT-1, TBP, GAPDH (left ventricle). The normalization strategy was tested analyzing mRNA expression of TNF-α, which is known to be up-regulated in HF and whose variations resulted more significant when normalized with the appropriately selected reference genes. The findings obtained in this study underline the importance to provide a set of reference genes to normalize mRNA expression in HF and control minipigs. The use of unvalidated reference genes can generate biased results because also their expression could be altered by the experimental conditions.

Keywords
Reference gene; RT-PCR; Heart failure; mRNA expression; TNF-α; GeNorm
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Selection of reference genes for normalization of real-time PCR data in minipig heart failure model and evaluation of TNF-α mRNA expression
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 153, Issues 3–4, 20 May 2011, Pages 92–99
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us