title: The effects of reference genes in qRT-PCR assays for determining the immune response of bovine cells (MDBK) infected with the Bovine Viral Diarrhea Virus 1 (BVDV-1).
abstract: The bovine viral diarrhea virus (BVDV) causes significant economic losses to the dairy industry worldwide, and understanding its infection mechanisms would be extremely useful in designing new and efficient treatments. Due to the limited number of specific antibodies against bovine proteins, differential gene expression analyses are vital for researching host immune responses to viral infection. qRT-PCR provides a sensitive platform to conduct such gene expression analyses, but suitable housekeeping genes are needed for accurate transcript normalization. The present study assessed nine reference genes in bovine kidney cells under conditions of BVDV-1 infection, incubation with pathogen-associated molecular patterns, and co-incubation with BAY117085, a pharmacological inhibitor of the NF-κB signaling pathway. Analyses of Ct values using the BestKeeper and Normfinder programs ranked CD81, RPL4, and GAPDH as the most reliable reference genes. This determination of a stable set of reference genes in this culture system will facilitate analyses of expression levels for genes of interest.
authors.affiliation: Facultad de Ciencias, Instituto de Bioquímica y Microbiología, Laboratorio de virología, Universidad Austral de Chile, Campus Isla Teja S/N, Valdivia, Chile.
authors.forename: Fernanda
authors.initials: F
authors.lastname: Fredericksen
authors.affiliation: Facultad de Ciencias, Instituto de Bioquímica y Microbiología, Laboratorio de virología, Universidad Austral de Chile, Campus Isla Teja S/N, Valdivia, Chile.
authors.forename: Fredy
authors.initials: F
authors.lastname: Delgado
authors.affiliation: Manuka, Ruta Puerto Octay Km. 30, Chile. Electronic address: info@manuka.cl.
authors.forename: Cristian
authors.initials: C
authors.lastname: Cabrera
authors.affiliation: Facultad de Ciencias, Instituto de Bioquímica y Microbiología, Laboratorio de virología, Universidad Austral de Chile, Campus Isla Teja S/N, Valdivia, Chile.
authors.forename: Alejandro
authors.initials: A
authors.lastname: Yáñez
authors.affiliation: Facultad de Ciencias, Instituto de Bioquímica y Microbiología, Laboratorio de virología, Universidad Austral de Chile, Campus Isla Teja S/N, Valdivia, Chile.
authors.forename: Carrasco
authors.initials: C
authors.lastname: Gonzalo
authors.affiliation: Facultad de Ciencias, Instituto de Bioquímica y Microbiología, Laboratorio de virología, Universidad Austral de Chile, Campus Isla Teja S/N, Valdivia, Chile.
authors.forename: Melina
authors.initials: M
authors.lastname: Villalba
authors.affiliation: Facultad de Ciencias, Instituto de Bioquímica y Microbiología, Laboratorio de virología, Universidad Austral de Chile, Campus Isla Teja S/N, Valdivia, Chile. Electronic address: volavarria@uach.cl.
authors.forename: Víctor H
authors.initials: VH
authors.lastname: Olavarría
mesh_terms: D000818:Animals; D002417:Cattle; D002460:Cell Line; D030241:Diarrhea Virus 1, Bovine Viral / Q000235:genetics* / Q000276:immunology / Q000472:pathogenicity; D004847:Epithelial Cells / Q000821:virology; D015967:Gene Expression Regulation, Viral / Q000235:genetics; D007111:Immunity, Cellular / Q000235:genetics*; D016328:NF-kappa B / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000235:genetics
chemical_list: D016328:NF-kappa B
keywords: BVDV-1; Flaviviridae; Housekeeping genes; Virus; qRT-PCR