ArticleInternational journal of obesity (2005)2014
Identification of optimal reference genes for RT-qPCR in the rat hypothalamus and intestine for the study of obesity.
Article in International journal of obesity (2005), 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 24 papers, 1 of them a synthesis that pooled it.
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Who cites it
24 citing papers in PubMed, 1 synthesis or guideline pooled it, 61 citations in OpenAlex.
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- Alanyl-glutamine protects the intestinal barrier function in trained rats against the impact of acute exhaustive exercise.Brazilian journal of medical and biological research = Revista brasileira de pesquisas medicas e biologica · 2020Article
- High-Fat Diet Alters the Expression of Reference Genes in Male Mice.Frontiers in nutrition · 2020Article
- Use of the REVERTMolecular and cellular endocrinology · 2018Article
- Evaluation of the performance of serum miRNAs as normalizers in microRNA studies focused on cardiovascular disease.Journal of thoracic disease · 2018Article
- Bioinformatics Analysis of Genes and Pathways of CD11bCurrent medical science · 2018Article
- Valid gene expression normalization by RT-qPCR in studies on hPDL fibroblasts with focus on orthodontic tooth movement and periodontitis.Scientific reports · 2017Article
- Validation of optimal reference genes for quantitative real time PCR in muscle and adipose tissue for obesity and diabetes research.Scientific reports · 2017Article
- Identification of suitable reference genes for real-time quantitative PCR analysis of hydrogen peroxide-treated human umbilical vein endothelial cells.BMC molecular biology · 2017Article
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Authors and funding
6 authors at 1 institution in 1 country.
Funding
Abstract
backgroundObesity has a complicated metabolic pathology, and defining the underlying mechanisms of obesity requires integrative studies with molecular end points. Real-time quantitative PCR (RT-qPCR) is a powerful tool that has been widely utilized. However, the importance of using carefully validated reference genes in RT-qPCR seems to have been overlooked in obesity-related research. The objective of this study was to select a set of reference genes with stable expressions to be used for RT-qPCR normalization in rats under fasted vs re-fed and chow vs high-fat diet (HFD) conditions.
designMale long-Evans rats were treated under four conditions: chow/fasted, chow/re-fed, HFD/fasted and HFD/re-fed. Expression stabilities of 13 candidate reference genes were evaluated in the rat hypothalamus, duodenum, jejunum and ileum using the ReFinder software program. The optimal number of reference genes needed for RT-qPCR analyses was determined using geNorm.
resultsUsing geNorm analysis, we found that it was sufficient to use the two most stably expressed genes as references in RT-qPCR analyses for each tissue under specific experimental conditions. B2M and RPLP0 in the hypothalamus, RPS18 and HMBS in the duodenum, RPLP2 and RPLP0 in the jejunum and RPS18 and YWHAZ in the ileum were the most suitable pairs for a normalization study when the four aforementioned experimental conditions were considered.
conclusionsOur study demonstrates that gene expression levels of reference genes commonly used in obesity-related studies, such as ACTB or RPS18, are altered by changes in acute or chronic energy status. These findings underline the importance of using reference genes that are stable in expression across experimental conditions when studying the rat hypothalamus and intestine, because these tissues have an integral role in the regulation of energy homeostasis. It is our hope that this study will raise awareness among obesity researchers on the essential need for reference gene validation in gene expression studies.
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