Validation of optimal reference genes for quantitative real time PCR in muscle and adipose tissue for obesity and diabetes research.
Sci Rep
; 7(1): 3612, 2017 06 15.
Article
en En
| MEDLINE
| ID: mdl-28620170
ABSTRACT
The global incidence of obesity has led to an increasing need for understanding the molecular mechanisms that drive this epidemic and its comorbidities. Quantitative real-time RT-PCR (RT-qPCR) is the most reliable and widely used method for gene expression analysis. The selection of suitable reference genes (RGs) is critical for obtaining accurate gene expression information. The current study aimed to identify optimal RGs to perform quantitative transcriptomic analysis based on RT-qPCR for obesity and diabetes research, employing in vitro and mouse models, and human tissue samples. Using the ReFinder program we evaluated the stability of a total of 15 RGs. The impact of choosing the most suitable RGs versus less suitable RGs on RT-qPCR results was assessed. Optimal RGs differed between tissue and cell type, species, and experimental conditions. By employing different sets of RGs to normalize the mRNA expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α), we show that sub-optimal RGs can markedly alter the PGC1α gene expression profile. Our study demonstrates the importance of validating RGs prior to normalizing transcriptional expression levels of target genes and identifies optimal RG pairs for reliable RT-qPCR normalization in cells and in human and murine muscle and adipose tissue for obesity/diabetes research.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Tejido Adiposo
/
Predisposición Genética a la Enfermedad
/
Diabetes Mellitus
/
Estudios de Asociación Genética
/
Músculos
/
Obesidad
Tipo de estudio:
Prognostic_studies
Límite:
Animals
Idioma:
En
Revista:
Sci Rep
Año:
2017
Tipo del documento:
Article
País de afiliación:
Canadá
Pais de publicación:
ENGLAND
/
ESCOCIA
/
GB
/
GREAT BRITAIN
/
INGLATERRA
/
REINO UNIDO
/
SCOTLAND
/
UK
/
UNITED KINGDOM