Senevirathna Jayan D M, Yonezawa Ryo, Saka Taiki, Igarashi Yoji, Funasaka Noriko, Yoshitake Kazutoshi, Kinoshita Shigeharu, Asakawa Shuichi
Laboratory of Aquatic Molecular Biology and Biotechnology Department of Aquatic Bioscience Graduate School of Agricultural and Life Sciences The University of Tokyo Tokyo Japan.
Department of Animal Science Faculty of Animal Science and Export Agriculture Uva Wellassa University Badulla Sri Lanka.
Ecol Evol. 2021 Nov 26;11(23):17142-17159. doi: 10.1002/ece3.8354. eCollection 2021 Dec.
Toothed whales are one group of marine mammals that has developed special adaptations, such as echolocation for predation, to successfully live in a dynamic aquatic environment. Their fat metabolism may differ from that of other mammals because toothed whales have acoustic fats. Gene expression in the metabolic pathways of animals can change with respect to their evolution and environment. A real-time quantitative polymerase chain reaction (RT-qPCR) is a reliable technique for studying the relative expressions of genes. However, since the accuracy of RT-qPCR data is totally dependent on the reference gene, the selection of the reference gene is an essential step. In this study, 10 candidate reference genes (, , , , , , , , , and ) were initially tested for amplification efficiency using RT-qPCR. After excluding , the remaining nine genes, which are nearly 100% efficient, were selected for the gene stability analysis. Stable reference genes across eight different fat tissue, liver, and muscle samples from were identified by four algorithms, which were provided in Genorm, NormFinder, BestKeeper, and Delta CT. Finally, a RefFinder comprehensive ranking was performed based on the stability values, and the nine genes were ranked as follows: > > > > > > > > . The and genes were identified as the most stable novel reference genes. The third-ranked gene, , is a well-known housekeeping gene for mammals. Ultimately, we suggest the use of as a reliable novel reference gene for genomics studies on the lipid-related aquatic adaptations of toothed whales.
齿鲸是一类海洋哺乳动物，它们已经进化出了特殊的适应性特征，比如用于捕食的回声定位，从而能够成功地生活在动态的水生环境中。它们的脂肪代谢可能与其他哺乳动物不同，因为齿鲸拥有声学脂肪。动物代谢途径中的基因表达会随着其进化和环境而发生变化。实时定量聚合酶链反应（RT-qPCR）是研究基因相对表达的可靠技术。然而，由于RT-qPCR数据的准确性完全依赖于参考基因，所以参考基因的选择是至关重要的一步。在本研究中，最初使用RT-qPCR对10个候选参考基因（、、、、、、、、和）进行了扩增效率测试。在排除后，选择了其余9个扩增效率接近100%的基因进行基因稳定性分析。通过Genorm、NormFinder、BestKeeper和Delta CT中提供的四种算法，在来自的八个不同脂肪组织、肝脏和肌肉样本中鉴定出了稳定的参考基因。最后，基于稳定性值进行了RefFinder综合排名，这9个基因的排名如下：> > > > > > > > 。和基因被确定为最稳定的新型参考基因。排名第三的基因是哺乳动物中一个著名的管家基因。最终，我们建议将作为齿鲸脂质相关水生适应性基因组学研究的可靠新型参考基因。
