Selection of reference genes for quantitative real-time PCR analysis in chicken embryo fibroblasts infected with avian leukosis virus subgroup J.

Abstract

BACKGROUND: The selection of stably expressed reference genes is a prerequisite when evaluating gene expression, via real-time PCR, in cells in response to viral infections. The objective of our study was to identify suitable reference genes for mRNA expression analysis in chicken embryonic fibroblasts (CEF) after infection with avian leukosis virus subgroup J (ALV-J). FINDINGS: The expression levels of 11 potential reference genes in CEF infected with ALV-J were determined by real-time PCR. The expression stability of these genes were analyzed and ranked using the geNorm tool. Analysis indicated that the genes RPL30 (ribosomal protein L30) and SDHA (succinate dehydrogenase complex, subunit A) were the most stably expressed genes in the ALV-J infected CEF. CONCLUSIONS: The RPL30 and SDHA were deemed suitable for use as reference genes for real-time PCR analysis of mRNA gene expression during ALV-J infection, whereas commonly used ACTB and GAPDH are unsuitable to be reference genes.