Evaluation of potential internal references for quantitative real-time RT-PCR normalization of gene expression in red drum (Sciaenops ocellatus)

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• 作者：Bo-guang Sun ; Yong-hua Hu
• 关键词：Housekeeping gene ; Expression stability ; Reference gene ; Sciaenops ocellatus
• 刊名：Fish Physiology and Biochemistry
• 出版年：2015
• 出版时间：June 2015
• 年：2015
• 卷：41
• 期：3
• 页码：695-704
• 全文大小：1,693 KB
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  Radonic A, Thulke S, Mackay IM, L
• 作者单位：Bo-guang Sun (1)
  Yong-hua Hu (1)
  
  1. Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao, 266071, People’s Republic of China
  
• 刊物类别：Biomedical and Life Sciences
• 刊物主题：Life Sciences
  Hydrobiology
  Animal Physiology
  Animal Anatomy, Morphology and Histology
  Animal Biochemistry
  Zoology
  
• 出版者：Springer Netherlands
• ISSN：1573-5168

文摘

Quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) has been used extensively for studying gene expression in diverse organisms including fish. In this study, with an aim to identify reliable reference genes for qRT-PCR in red drum (Sciaenops ocellatus), an economic fish species, we determined the expression stability of seven housekeeping genes in healthy and bacterium-infected red drum. Each of the selected candidate genes was amplified by qRT-PCR from the brain, gill, heart, intestine, kidney, liver, muscle, and spleen of red drum infected with or without a bacterial pathogen for 12 and 48?h. The mRNA levels of the genes were analyzed with the geNorm and NormFinder algorithms. The results showed that in the absence of bacterial infection, translation initiation factor 3, NADH dehydrogenase 1, and QM-like protein may be used together as internal references across the eight examined tissues. Bacterial infection caused variations in the rankings of the most stable genes in a tissue-dependent manner. For all tissues, two genes sufficed for reliable normalization at both 12 and 48?h post-infection. However, the optimal gene pairs differed among tissues and, for four of the examined eight tissues, between infection points. These results indicate that when studying gene expression in red drum under conditions of bacterial infection, the optimal reference genes should be selected on the basis of tissue type and, for accurate normalization, infection stage.