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Variation in Stability of Endogenous Reference Genes in Fallopian Tubes and Endometrium from Healthy and Ectopic Pregnant Women

RT-qPCR is commonly employed in gene expression studies in ectopic pregnancy. Most use RN18S1, β-actin or GAPDH as internal controls without validation of their suitability as reference genes. A systematic study of the suitability of endogenous reference genes for gene expression studies in ectopic...

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Autores principales: Gebeh, Alpha K., Marczylo, Emma L., Amoako, Akwasi A., Willets, Jonathon M., Konje, Justin C.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Molecular Diversity Preservation International (MDPI) 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3317689/
https://www.ncbi.nlm.nih.gov/pubmed/22489127
http://dx.doi.org/10.3390/ijms13032810
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author Gebeh, Alpha K.
Marczylo, Emma L.
Amoako, Akwasi A.
Willets, Jonathon M.
Konje, Justin C.
author_facet Gebeh, Alpha K.
Marczylo, Emma L.
Amoako, Akwasi A.
Willets, Jonathon M.
Konje, Justin C.
author_sort Gebeh, Alpha K.
collection PubMed
description RT-qPCR is commonly employed in gene expression studies in ectopic pregnancy. Most use RN18S1, β-actin or GAPDH as internal controls without validation of their suitability as reference genes. A systematic study of the suitability of endogenous reference genes for gene expression studies in ectopic pregnancy is lacking. The aims of this study were therefore to evaluate the stability of 12 reference genes and suggest those that are stable for use as internal control genes in fallopian tubes and endometrium from ectopic pregnancy and healthy non-pregnant controls. Analysis of the results showed that the genes consistently ranked in the top six by geNorm and NormFinder algorithms, were UBC, GAPDH, CYC1 and EIF4A2 (fallopian tubes) and UBC and ATP5B (endometrium). mRNA expression of NAPE-PLD as a test gene of interest varied between the groups depending on which of the 12 reference genes was used as internal controls. This study demonstrates that arbitrary selection of reference genes for normalisation in RT-qPCR studies in ectopic pregnancy without validation, risk producing inaccurate data and should therefore be discouraged.
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spelling pubmed-33176892012-04-09 Variation in Stability of Endogenous Reference Genes in Fallopian Tubes and Endometrium from Healthy and Ectopic Pregnant Women Gebeh, Alpha K. Marczylo, Emma L. Amoako, Akwasi A. Willets, Jonathon M. Konje, Justin C. Int J Mol Sci Article RT-qPCR is commonly employed in gene expression studies in ectopic pregnancy. Most use RN18S1, β-actin or GAPDH as internal controls without validation of their suitability as reference genes. A systematic study of the suitability of endogenous reference genes for gene expression studies in ectopic pregnancy is lacking. The aims of this study were therefore to evaluate the stability of 12 reference genes and suggest those that are stable for use as internal control genes in fallopian tubes and endometrium from ectopic pregnancy and healthy non-pregnant controls. Analysis of the results showed that the genes consistently ranked in the top six by geNorm and NormFinder algorithms, were UBC, GAPDH, CYC1 and EIF4A2 (fallopian tubes) and UBC and ATP5B (endometrium). mRNA expression of NAPE-PLD as a test gene of interest varied between the groups depending on which of the 12 reference genes was used as internal controls. This study demonstrates that arbitrary selection of reference genes for normalisation in RT-qPCR studies in ectopic pregnancy without validation, risk producing inaccurate data and should therefore be discouraged. Molecular Diversity Preservation International (MDPI) 2012-03-02 /pmc/articles/PMC3317689/ /pubmed/22489127 http://dx.doi.org/10.3390/ijms13032810 Text en © 2012 by the authors; licensee Molecular Diversity Preservation International, Basel, Switzerland. http://creativecommons.org/licenses/by/3.0 This article is an open-access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/3.0/).
spellingShingle Article
Gebeh, Alpha K.
Marczylo, Emma L.
Amoako, Akwasi A.
Willets, Jonathon M.
Konje, Justin C.
Variation in Stability of Endogenous Reference Genes in Fallopian Tubes and Endometrium from Healthy and Ectopic Pregnant Women
title Variation in Stability of Endogenous Reference Genes in Fallopian Tubes and Endometrium from Healthy and Ectopic Pregnant Women
title_full Variation in Stability of Endogenous Reference Genes in Fallopian Tubes and Endometrium from Healthy and Ectopic Pregnant Women
title_fullStr Variation in Stability of Endogenous Reference Genes in Fallopian Tubes and Endometrium from Healthy and Ectopic Pregnant Women
title_full_unstemmed Variation in Stability of Endogenous Reference Genes in Fallopian Tubes and Endometrium from Healthy and Ectopic Pregnant Women
title_short Variation in Stability of Endogenous Reference Genes in Fallopian Tubes and Endometrium from Healthy and Ectopic Pregnant Women
title_sort variation in stability of endogenous reference genes in fallopian tubes and endometrium from healthy and ectopic pregnant women
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3317689/
https://www.ncbi.nlm.nih.gov/pubmed/22489127
http://dx.doi.org/10.3390/ijms13032810
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