An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either Piscirickettsia salmonis or IPNV

Peña, Andrea A; Bols, Niels C; Marshall, Sergio H

physiological states and to play a role in a broad range of cellular mechanisms [29] . GAPDH expression was increasingly upregulated in RTS11 cells infected with P. salmonis and was slightly upregulated over 48 h in both control and IPNV-treated CHSE-214 cultures. This regulation on non-infected cells was not perceptible in the long-term experiment with P. salmonis or in the assays with RTS11. Schmitten and Zakrajsek [30] demonstrated that the addition of serum to serum-starved human fibroblast cultures increased the mRNA amount for GAPDH. Although in our experiments, cells were maintained in a low serum concentration (5% fetal bovine serum), the addition of fresh media prior to infection could stimulate slightly the expression of GAPDH in the first few hours. Therefore, an important point to consider during the selection of a reference gene is whether expression could be eventually regulated by other factors than the challenge self. This differential regulation in the reference gene could lead to over-or sub-estimations of target gene expression. As well, TUBA showed increased expression over 48 h in both control and IPNV-infected CHSE-214 cultures, while in infected RTS11 cultures, TUBA was downregulated. Then, TUBA gene expression was influenced by IPNV infection but the direction of the influence depended on the cell type.