doiSerbia

Selection of reference genes for normalization of real-time PCR data in visceral adipose tissue of female rats on a fructose-enriched diet

Elaković Ivana (Institute for Biological Research “Siniša Stanković”, Department of Biochemistry, Belgrade)
Nestorov Jelena (Institute for Biological Research “Siniša Stanković”, Department of Biochemistry, Belgrade)
Kovačević Sanja (Institute for Biological Research “Siniša Stanković”, Department of Biochemistry, Belgrade)
Matić Gordana (Institute for Biological Research “Siniša Stanković”, Department of Biochemistry, Belgrade)

Considering the association of increased fructose intake with the rise in metabolic syndrome and gender differences in its incidence, this study was aimed at evaluating and identifying the optimal reference genes for the relative quantification of gene expression by real-time PCR in the adipose tissue of female rats exposed to a fructose-enriched diet as an animal model of metabolic syndrome. The transcription of four commonly used reference genes, BA, HPRT1, TBP and B2M, was quantified by TaqMan real-time PCR. The expression stability of these genes was evaluated by direct comparison of Ct values and by using the geNorm and NormFinder algorithms. NormFinder revealed HPRT1 as the most stable gene, and both programs identified HPRT1 and TBP as the most stable combination of genes in this experimental paradigm. In addition, HPRT1 was shown to be reliable enough as the sole reference gene.

Keywords: Reference genes, real-time PCR, fructose diet, adipose tissue, female rats

Projekat Ministarstva nauke Republike Srbije, br. III41009