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

Tissue-specific selection of reference genes is required for expression studies in the mouse model of maternal protein undernutrition
Tissue-specific selection of reference genes is required for expression studies in the mouse model of maternal protein undernutrition
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.
reference genes, gene expression, real-time qPCR, mouse embryos, mouse embryonic stem cells, maternal diet
0093-691X
558-569
Lucas, E.S.
4f46398a-5f53-4e61-875e-40b5013b1d2d
Watkins, A.J.
106e91fc-632c-4a7b-b412-30118a75ce4e
Cox, A.L.
e9f28fdf-70a7-4278-bd83-774284244799
Marfy-Smith, S.J.
2020fb0a-d6eb-4516-b390-308bba053de9
Smyth, N.
0eba2a40-3b43-4d40-bb64-621bd7e9d505
Fleming, T.P.
2abf761a-e5a1-4fa7-a2c8-12e32d5d4c03
Lucas, E.S.
4f46398a-5f53-4e61-875e-40b5013b1d2d
Watkins, A.J.
106e91fc-632c-4a7b-b412-30118a75ce4e
Cox, A.L.
e9f28fdf-70a7-4278-bd83-774284244799
Marfy-Smith, S.J.
2020fb0a-d6eb-4516-b390-308bba053de9
Smyth, N.
0eba2a40-3b43-4d40-bb64-621bd7e9d505
Fleming, T.P.
2abf761a-e5a1-4fa7-a2c8-12e32d5d4c03

Lucas, E.S., Watkins, A.J., Cox, A.L., Marfy-Smith, S.J., Smyth, N. and Fleming, T.P. (2011) Tissue-specific selection of reference genes is required for expression studies in the mouse model of maternal protein undernutrition. Theriogenology, 76 (3), 558-569. (doi:10.1016/j.theriogenology.2011.03.008). (PMID:21550104)

Record type: Article

Abstract

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.

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More information

e-pub ahead of print date: 7 May 2011
Published date: August 2011
Keywords: reference genes, gene expression, real-time qPCR, mouse embryos, mouse embryonic stem cells, maternal diet
Organisations: Biomedicine

Identifiers

Local EPrints ID: 343321
URI: https://eprints.soton.ac.uk/id/eprint/343321
ISSN: 0093-691X
PURE UUID: 3b776b60-886f-4792-b727-33bd279bba6a

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Date deposited: 03 Oct 2012 10:32
Last modified: 16 Jul 2019 21:53

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