Abstract
BackgroundSalmonid fishes are among the most widely studied model fish species but reports on systematic evaluation of reference genes in qRT-PCR studies is lacking.ResultsThe stability of six potential reference genes was examined in eight tissues of Atlantic salmon (Salmo salar), to determine the most suitable genes to be used in quantitative real-time RT-PCR analyses. The relative transcription levels of genes encoding 18S rRNA, S20 ribosomal protein, β-actin, glyceraldehyde-3P-dehydrogenase (GAPDH), and two paralog genes encoding elongation factor 1A (EF1AA and EF1AB) were quantified in gills, liver, head kidney, spleen, thymus, brain, muscle, and posterior intestine in six untreated adult fish, in addition to a group of individuals that went through smoltification. Based on calculations performed with the geNorm VBA applet, which determines the most stable genes from a set of tested genes in a given cDNA sample, the ranking of the examined genes in adult Atlantic salmon was EF1AB>EF1AA>β-actin>18S rRNA>S20>GAPDH. When the same calculations were done on a total of 24 individuals from four stages in the smoltification process (presmolt, smolt, smoltified seawater and desmoltified freshwater), the gene ranking was EF1AB>EF1AA>S20>β-actin>18S rRNA>GAPDH.ConclusionOverall, this work suggests that the EF1AA and EF1AB genes can be useful as reference genes in qRT-PCR examination of gene expression in the Atlantic salmon.
Highlights
Salmonid fishes are among the most widely studied model fish species but reports on systematic evaluation of reference genes in qRT-PCR studies is lacking
Ribosomal RNA and total RNA are most commonly used as references in gene expression analysis today
Our data, based on geNorm calculations, suggest that the Atlantic salmon EF1A genes that have been tested in the present study may be good candidate reference genes
Summary
Salmonid fishes are among the most widely studied model fish species but reports on systematic evaluation of reference genes in qRT-PCR studies is lacking. Ribosomal RNA (rRNA) and total RNA are most commonly used as references in gene expression analysis today. The purpose of these controls is to remove or reduce differences due to sampling, i.e. differences in RNA quantity and quality. The ideal endogenous control should be expressed at a constant level among different tissues of an organism, at all stages of development and should be unaffected by the experimental treatment. It should be expressed at roughly the same level as the RNA under study [1]. Numerous studies have revealed that no single universal (page number not for citation purposes)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
