Evaluation of qPCR reference genes in GH-overexpressing transgenic zebrafish (Danio rerio)
| Autor: | Amanda Weege Da Silveira Martins, Tony Silveira, Larissa O. Daneluz, Eliza Rossi Komninou, William Borges Domingues, Hadassa G. Ortiz, Mariana H. Remião, Gabriela T. Rassier, Luis Fernando Marins, Mateus Tavares Kütter, Eduardo N. Dellagostin, Vinicius Farias Campos |
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| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Transgene Danio lcsh:Medicine Real-Time Polymerase Chain Reaction Article Animals Genetically Modified 03 medical and health sciences Gene expression analysis 0302 clinical medicine Reference genes Gene expression Animals lcsh:Science Muscle Skeletal Zebrafish Gene Glyceraldehyde 3-phosphate dehydrogenase Brain Chemistry Multidisciplinary biology lcsh:R fungi Reference Standards Zebrafish Proteins biology.organism_classification Eukaryotic translation elongation factor 1 alpha 1 Cell biology Intestines 030104 developmental biology Liver Growth Hormone Genetic engineering Reverse transcription polymerase chain reaction biology.protein lcsh:Q Software 030217 neurology & neurosurgery |
| Zdroj: | Scientific Reports Scientific Reports, Vol 10, Iss 1, Pp 1-15 (2020) |
| ISSN: | 2045-2322 |
| Popis: | Reference genes (RGs) must have a stable expression in tissues in all experimental conditions to normalize real-time quantitative reverse transcription PCR (qRT-PCR) data. F0104 is a highly studied lineage of zebrafish developed to overexpress the growth hormone (GH). It is assumed that the transgenic process may influence the expression levels of commonly used RGs. The objective of the present study was to make a comprehensive analysis of stability of canditade RGs actb1, actb2, b2m, eif2s2, eef1a1, gapdh, rplp2, rpl7, rpl13α, tuba1, and rps18, in gh-transgenic and non-transgenic zebrafish. Liver, brain, intestine and muscle samples from both groups had qRT-PCR results analyzed by dCt, geNorm, NormFinder, BestKeeper, and RefFinder softwares. Consensus analyses among software concluded that rpl13α, rpl7, and eef1a1 are the most stable genes for zebrafish, considering the studied groups and tissues. Gapdh, rps18, and tuba1 suffered variations in stability among different tissues of both groups, and so, they were listed as the genes with lowest stability. Results from an average pairwise variations test indicated that the use of two RGs would generate reliable results for gene expression analysis in the studied tissues. We conclude that genes that are commonly used in mammals for qRT-PCR assays have low stability in both non-transgenic and gh-transgenic zebrafish reinforcing the importance of using species-specific RGs. |
| Databáze: | OpenAIRE |
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