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A probe-based qRT-PCR method to profile immunological gene expression in blood of captive beluga whales (Delphinapterus leucas)

Cytokines are fundamental for a functioning immune system, and thus potentially serve as important indicators of animal health. Quantitation of mRNA using quantitative reverse transcription polymerase chain reaction (qRT-PCR) is an established immunological technique. It is particularly suitable for...

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Autores principales: Tsai, Ming-An, Chen, I-Hua, Wang, Jiann-Hsiung, Chou, Shih-Jen, Li, Tsung-Hsien, Leu, Ming-Yih, Ho, Hsiao-Kuan, Yang, Wei Cheng
Formato: Online Artículo Texto
Lenguaje:English
Publicado: PeerJ Inc. 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5622604/
https://www.ncbi.nlm.nih.gov/pubmed/28970970
http://dx.doi.org/10.7717/peerj.3840
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author Tsai, Ming-An
Chen, I-Hua
Wang, Jiann-Hsiung
Chou, Shih-Jen
Li, Tsung-Hsien
Leu, Ming-Yih
Ho, Hsiao-Kuan
Yang, Wei Cheng
author_facet Tsai, Ming-An
Chen, I-Hua
Wang, Jiann-Hsiung
Chou, Shih-Jen
Li, Tsung-Hsien
Leu, Ming-Yih
Ho, Hsiao-Kuan
Yang, Wei Cheng
author_sort Tsai, Ming-An
collection PubMed
description Cytokines are fundamental for a functioning immune system, and thus potentially serve as important indicators of animal health. Quantitation of mRNA using quantitative reverse transcription polymerase chain reaction (qRT-PCR) is an established immunological technique. It is particularly suitable for detecting the expression of proteins against which monoclonal antibodies are not available. In this study, we developed a probe-based quantitative gene expression assay for immunological assessment of captive beluga whales (Delphinapterus leucas) that is one of the most common cetacean species on display in aquariums worldwide. Six immunologically relevant genes (IL-2Rα, -4, -10, -12, TNFα, and IFNγ) were selected for analysis, and two validated housekeeping genes (PGK1 and RPL4) with stable expression were used as reference genes. Sixteen blood samples were obtained from four animals with different health conditions and stored in RNAlater™ solution. These samples were used for RNA extraction followed by qRT-PCR analysis. Analysis of gene transcripts was performed by relative quantitation using the comparative Cq method with the integration of amplification efficiency and two reference genes. The expression levels of each gene in the samples from clinically healthy animals were normally distributed. Transcript outliers for IL-2Rα, IL-4, IL-12, TNFα, and IFNγ were noticed in four samples collected from two clinically unhealthy animals. This assay has the potential to identify immune system deviation from normal state, which is caused by health problems. Furthermore, knowing the immune status of captive cetaceans could help both trainers and veterinarians in implementing preventive approaches prior to disease onset.
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spelling pubmed-56226042017-10-02 A probe-based qRT-PCR method to profile immunological gene expression in blood of captive beluga whales (Delphinapterus leucas) Tsai, Ming-An Chen, I-Hua Wang, Jiann-Hsiung Chou, Shih-Jen Li, Tsung-Hsien Leu, Ming-Yih Ho, Hsiao-Kuan Yang, Wei Cheng PeerJ Marine Biology Cytokines are fundamental for a functioning immune system, and thus potentially serve as important indicators of animal health. Quantitation of mRNA using quantitative reverse transcription polymerase chain reaction (qRT-PCR) is an established immunological technique. It is particularly suitable for detecting the expression of proteins against which monoclonal antibodies are not available. In this study, we developed a probe-based quantitative gene expression assay for immunological assessment of captive beluga whales (Delphinapterus leucas) that is one of the most common cetacean species on display in aquariums worldwide. Six immunologically relevant genes (IL-2Rα, -4, -10, -12, TNFα, and IFNγ) were selected for analysis, and two validated housekeeping genes (PGK1 and RPL4) with stable expression were used as reference genes. Sixteen blood samples were obtained from four animals with different health conditions and stored in RNAlater™ solution. These samples were used for RNA extraction followed by qRT-PCR analysis. Analysis of gene transcripts was performed by relative quantitation using the comparative Cq method with the integration of amplification efficiency and two reference genes. The expression levels of each gene in the samples from clinically healthy animals were normally distributed. Transcript outliers for IL-2Rα, IL-4, IL-12, TNFα, and IFNγ were noticed in four samples collected from two clinically unhealthy animals. This assay has the potential to identify immune system deviation from normal state, which is caused by health problems. Furthermore, knowing the immune status of captive cetaceans could help both trainers and veterinarians in implementing preventive approaches prior to disease onset. PeerJ Inc. 2017-09-27 /pmc/articles/PMC5622604/ /pubmed/28970970 http://dx.doi.org/10.7717/peerj.3840 Text en ©2017 Tsai 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 (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited.
spellingShingle Marine Biology
Tsai, Ming-An
Chen, I-Hua
Wang, Jiann-Hsiung
Chou, Shih-Jen
Li, Tsung-Hsien
Leu, Ming-Yih
Ho, Hsiao-Kuan
Yang, Wei Cheng
A probe-based qRT-PCR method to profile immunological gene expression in blood of captive beluga whales (Delphinapterus leucas)
title A probe-based qRT-PCR method to profile immunological gene expression in blood of captive beluga whales (Delphinapterus leucas)
title_full A probe-based qRT-PCR method to profile immunological gene expression in blood of captive beluga whales (Delphinapterus leucas)
title_fullStr A probe-based qRT-PCR method to profile immunological gene expression in blood of captive beluga whales (Delphinapterus leucas)
title_full_unstemmed A probe-based qRT-PCR method to profile immunological gene expression in blood of captive beluga whales (Delphinapterus leucas)
title_short A probe-based qRT-PCR method to profile immunological gene expression in blood of captive beluga whales (Delphinapterus leucas)
title_sort probe-based qrt-pcr method to profile immunological gene expression in blood of captive beluga whales (delphinapterus leucas)
topic Marine Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5622604/
https://www.ncbi.nlm.nih.gov/pubmed/28970970
http://dx.doi.org/10.7717/peerj.3840
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