A novel universal probe library quantitative reverse transcription polymerase chain reaction method to profile immunological gene expression in blood of captive beluga whales (Delphinapterusleucas)
- Published
- Accepted
- Subject Areas
- Marine Biology, Molecular Biology, Veterinary Medicine, Immunology
- Keywords
- Gene expression, qRT-PCR, Immunology, Beluga
- Copyright
- © 2017 Tsai et al.
- Licence
- This is an open access article distributed under the terms of the Creative Commons Attribution License, 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 Preprints) and either DOI or URL of the article must be cited.
- Cite this article
- 2017. A novel universal probe library quantitative reverse transcription polymerase chain reaction method to profile immunological gene expression in blood of captive beluga whales (Delphinapterusleucas) PeerJ Preprints 5:e2735v1 https://doi.org/10.7287/peerj.preprints.2735v1
Abstract
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 novel probe-based quantitative gene expression assay for immunological assessment of captive beluga whales ( Delphinapterusleucas ) 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.
Author Comment
This is a submission to PeerJ for review.
Supplemental Information
Cq values of the tested samples
Cq values of 6 target genes and 2 reference genes of the 12 healthy samples and 4 non-healthy samples.