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Identification of Reference Genes for Relative Quantification of Circulating MicroRNAs in Bovine Serum

Circulating microRNAs in body fluids have been implicated as promising biomarkers for physiopathology disorders. Currently, the expression levels of circulating microRNAs are estimated by reverse transcription quantitative real-time polymerase chain reaction. Use of appropriate reference microRNAs f...

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Autores principales: Bae, In-Seon, Chung, Ki Yong, Yi, Jongmin, Kim, Tae Il, Choi, Hwa-Sik, Cho, Young-Moo, Choi, Inho, Kim, Sang Hoon
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4380332/
https://www.ncbi.nlm.nih.gov/pubmed/25826387
http://dx.doi.org/10.1371/journal.pone.0122554
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author Bae, In-Seon
Chung, Ki Yong
Yi, Jongmin
Kim, Tae Il
Choi, Hwa-Sik
Cho, Young-Moo
Choi, Inho
Kim, Sang Hoon
author_facet Bae, In-Seon
Chung, Ki Yong
Yi, Jongmin
Kim, Tae Il
Choi, Hwa-Sik
Cho, Young-Moo
Choi, Inho
Kim, Sang Hoon
author_sort Bae, In-Seon
collection PubMed
description Circulating microRNAs in body fluids have been implicated as promising biomarkers for physiopathology disorders. Currently, the expression levels of circulating microRNAs are estimated by reverse transcription quantitative real-time polymerase chain reaction. Use of appropriate reference microRNAs for normalization is critical for accurate microRNA expression analysis. However, no study has systematically investigated reference genes for evaluating circulating microRNA expression in cattle. In this study, we describe the identification and characterization of appropriate reference microRNAs for use in the normalization of circulating microRNA levels in bovine serum. We evaluated the expression stability of ten candidate reference genes in bovine serum by using reverse transcription quantitative real-time polymerase chain reaction. Data were analyzed using geNorm, NormFinder, and BestKeeper statistical algorithms. The results consistently showed that a combination of miR-93 and miR-127 provided the most stably expressed reference. The suitability of these microRNAs was validated, and even when compared among different genders or breeds, the combination of miR-93 and miR-127 was ranked as the most stable microRNA reference. Therefore, we conclude that this combination is the optimal endogenous reference for reverse transcription quantitative real-time polymerase chain reaction-based detection of microRNAs in bovine serum. The data presented in this study are crucial to successful biomarker discovery and validation for the diagnosis of physiopathological conditions in cattle.
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spelling pubmed-43803322015-04-09 Identification of Reference Genes for Relative Quantification of Circulating MicroRNAs in Bovine Serum Bae, In-Seon Chung, Ki Yong Yi, Jongmin Kim, Tae Il Choi, Hwa-Sik Cho, Young-Moo Choi, Inho Kim, Sang Hoon PLoS One Research Article Circulating microRNAs in body fluids have been implicated as promising biomarkers for physiopathology disorders. Currently, the expression levels of circulating microRNAs are estimated by reverse transcription quantitative real-time polymerase chain reaction. Use of appropriate reference microRNAs for normalization is critical for accurate microRNA expression analysis. However, no study has systematically investigated reference genes for evaluating circulating microRNA expression in cattle. In this study, we describe the identification and characterization of appropriate reference microRNAs for use in the normalization of circulating microRNA levels in bovine serum. We evaluated the expression stability of ten candidate reference genes in bovine serum by using reverse transcription quantitative real-time polymerase chain reaction. Data were analyzed using geNorm, NormFinder, and BestKeeper statistical algorithms. The results consistently showed that a combination of miR-93 and miR-127 provided the most stably expressed reference. The suitability of these microRNAs was validated, and even when compared among different genders or breeds, the combination of miR-93 and miR-127 was ranked as the most stable microRNA reference. Therefore, we conclude that this combination is the optimal endogenous reference for reverse transcription quantitative real-time polymerase chain reaction-based detection of microRNAs in bovine serum. The data presented in this study are crucial to successful biomarker discovery and validation for the diagnosis of physiopathological conditions in cattle. Public Library of Science 2015-03-31 /pmc/articles/PMC4380332/ /pubmed/25826387 http://dx.doi.org/10.1371/journal.pone.0122554 Text en © 2015 Bae et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Bae, In-Seon
Chung, Ki Yong
Yi, Jongmin
Kim, Tae Il
Choi, Hwa-Sik
Cho, Young-Moo
Choi, Inho
Kim, Sang Hoon
Identification of Reference Genes for Relative Quantification of Circulating MicroRNAs in Bovine Serum
title Identification of Reference Genes for Relative Quantification of Circulating MicroRNAs in Bovine Serum
title_full Identification of Reference Genes for Relative Quantification of Circulating MicroRNAs in Bovine Serum
title_fullStr Identification of Reference Genes for Relative Quantification of Circulating MicroRNAs in Bovine Serum
title_full_unstemmed Identification of Reference Genes for Relative Quantification of Circulating MicroRNAs in Bovine Serum
title_short Identification of Reference Genes for Relative Quantification of Circulating MicroRNAs in Bovine Serum
title_sort identification of reference genes for relative quantification of circulating micrornas in bovine serum
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4380332/
https://www.ncbi.nlm.nih.gov/pubmed/25826387
http://dx.doi.org/10.1371/journal.pone.0122554
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