Tissue-specific selection of reference genes is required for expression studies in the mouse model of maternal protein undernutrition

• Published in: Theriogenology Vol. 76; no. 3; pp. 558 - 569
• Main Authors: Lucas, E.S., Watkins, A.J., Cox, A.L., Marfy-Smith, S.J., Smyth, N., Fleming, T.P.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.08.2011; [Oxford]: Butterworth-Heinemann; [New York]: Elsevier Science
• Subjects: animal models; Animals; blastocyst; Blastocyst - metabolism; embryonic stem cells; Embryonic Stem Cells - metabolism; Female; Gene Expression; Gene Expression Profiling; Gene Expression Regulation, Developmental; genes; heart; Intra-Abdominal Fat - metabolism; Kidney - metabolism; kidneys; liver; Liver - metabolism; malnutrition; Maternal diet; maternal nutrition; Maternal Nutritional Physiological Phenomena; metabolism; Mice; Mouse embryonic stem cells; Mouse embryos; Myocardium - metabolism; placenta; Placenta - metabolism; Pregnancy; progeny; Protein-Energy Malnutrition - genetics; Protein-Energy Malnutrition - metabolism; Real-time qPCR; Reference genes; Reference Values; risk; Sex Factors; yolk sac; Yolk Sac - metabolism; Mouse embryos; Real-time qPCR; Gene expression; Reference genes; Mouse embryonic stem cells; Maternal diet
• ISSN: 0093-691X; 1879-3231
• DOI: 10.1016/j.theriogenology.2011.03.008

Suboptimal maternal nutrition during gestation results in the establishment of long-term phenotypic changes and an increased disease risk in the offspring. To elucidate how such environmental sensitivity results in physiological outcomes, the molecular characterisation of these offspring has become the focus of many studies. However, the likely modification of key cellular processes such as metabolism in response to maternal undernutrition raises the question of whether the genes typically used as reference constants in gene expression studies are suitable controls. Using a mouse model of maternal protein undernutrition, we have investigated the stability of seven commonly used reference genes ( 18s, Hprt1, Pgk1, Ppib, Sdha, Tbp and Tuba1) in a variety of offspring tissues including liver, kidney, heart, retro-peritoneal and inter-scapular fat, extra-embryonic placenta and yolk sac, as well as in the preimplantation blastocyst and blastocyst-derived embryonic stem cells. We find that although the selected reference genes are all highly stable within this system, they show tissue, treatment and sex-specific variation. Furthermore, software-based selection approaches rank reference genes differently and do not always identify genes which differ between conditions. Therefore, we recommend that reference gene selection for gene expression studies should be thoroughly validated for each tissue of interest.