Hypoxanthine Guanine Phosphoribosyl Transferase Is the Most Stable Reference Gene for Gene Expression Analysis by Quantitative PCR in Peripheral Blood Mononuclear Cells from Women with the Polycystic Ovary Syndrome
2014
Аутори:
Vojnović-Milutinović, DanijelaMacut, Djuro
Bozic-Antic, Ivana
Macut, Jelica Bjekic
Radovanović, Marina
Matić, Gordana
Brkljačić, Jelena
Тип документа:
Чланак у часопису (Објављена верзија)
Метаподаци
Приказ свих података о документуАпстракт:
Background: The polycystic ovary syndrome (PCOS) is a frequent endocrine
disorder that affects women of reproductive age. As the syndrome is
strongly associated with obesity, it is of interest to examine the gene
expression differences that accompany its development and the associated
metabolic disturbances. Real-time RT PCR is a standard method for
studying changes in gene expression. However, to obtain accurate and
reliable results, validation of reference genes is obligatory. The aim
of this study was to identify a suitable reference for the normalization
of gene expression in peripheral blood mononuclear cells (PBMCs) from
obese and normal-weight women with PCOS.
Methods: The expression stability of four potential reference genes:
hypoxanthine guanine phosphoribosyl transferase 1 (HPRT), beta-actin
(BA), beta(2)-microglobulin (B2M) and glyceraldehyde-3-phosphate
dehydrogenase (GAPDH), was assessed in PBMCs from healthy women, and
from normal-weight and obese women with PCOS. The variability in the
expression of potential reference genes was analyzed by the TaqMan
real-time RT PCR method, using GeNorm and Norm Finder software packages.
Results: Direct comparison of cycle threshold (Ct) values showed
inter-individual variations for all validated genes, the Ct values of
HPRT being less variable than those of BA, GAPDH and B2M. Both software
packages pointed to HPRT as the most steadily expressed gene in the
PBMCs of women with PCOS and healthy controls.
Conclusions: Cross-validation of the expression stability of four
potential reference genes identified HPRT as the most stable reference,
suitable for further investigations of gene expression in PBMCs from
women with PCOS.
Кључне речи:
peripheral blood mononuclear cells; real-time RT PCR; reference geneИзвор:
Journal of Medical Biochemistry, 2014, 33, 4, 356-363Финансирање / пројекти:
- Улога стероидних хормона у неуроендокриној адаптацији на стрес и патофизиологији метаболичког синдрома - молекуларни механизми и клиничке импликације (RS-MESTD-Integrated and Interdisciplinary Research (IIR or III)-41009)
DOI: 10.2478/jomb-2014-0029
ISSN: 1452-8266