title: Selection of suitable reference genes for normalization of genes of interest in canine soft tissue sarcomas using quantitative real-time polymerase chain reaction.
abstract: Quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR) is a sensitive technique for quantifying gene expression. Stably expressed reference genes are necessary for normalization of RT-qPCR data. Only a few articles have been published on reference genes in canine tumours. The objective of this study was to demonstrate how to identify suitable reference genes for normalization of genes of interest in canine soft tissue sarcomas using RT-qPCR. Primer pairs for 17 potential reference genes were designed and tested in archival tumour biopsies from six dogs. The geNorm algorithm was used to analyse the most suitable reference genes. Eight potential reference genes were excluded from this final analysis because of their dissociation curves. β-Glucuronidase (GUSB) and proteasome subunit, beta type, 6 (PSMB6) were most stably expressed with an M value of 0.154 and a CV of 0.053 describing their average stability. We suggest that choice of reference genes should be based on specific testing in every new experimental set-up.
authors.affiliation: Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Frederiksberg C, Denmark.|Department of Clinical Physiology, Nuclear Medicine & PET, Copenhagen University Hospital and Cluster for Molecular Imaging, Faculty of Health Sciences, University of Copenhagen, Copenhagen N, Denmark.
authors.forename: K W
authors.initials: KW
authors.lastname: Zornhagen
authors.affiliation: Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Frederiksberg C, Denmark.
authors.forename: A T
authors.initials: AT
authors.lastname: Kristensen
authors.affiliation: Department of Clinical Physiology, Nuclear Medicine & PET, Copenhagen University Hospital and Cluster for Molecular Imaging, Faculty of Health Sciences, University of Copenhagen, Copenhagen N, Denmark.|Department of Micro- and Nanotechnology, Technical University of Denmark, Kgs. Lyngby, Denmark.
authors.forename: A E
authors.initials: AE
authors.lastname: Hansen
authors.affiliation: Department of Clinical Physiology, Nuclear Medicine & PET, Copenhagen University Hospital and Cluster for Molecular Imaging, Faculty of Health Sciences, University of Copenhagen, Copenhagen N, Denmark.
authors.forename: J
authors.initials: J
authors.lastname: Oxboel
authors.affiliation: Department of Clinical Physiology, Nuclear Medicine & PET, Copenhagen University Hospital and Cluster for Molecular Imaging, Faculty of Health Sciences, University of Copenhagen, Copenhagen N, Denmark.
authors.forename: A
authors.initials: A
authors.lastname: Kjaer
mesh_terms: D000818:Animals; D004283:Dog Diseases / Q000235:genetics*; D004285:Dogs / Q000235:genetics; D052138:Genes, Neoplasm / Q000235:genetics*; D005966:Glucuronidase / Q000235:genetics; D046988:Proteasome Endopeptidase Complex / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D012509:Sarcoma / Q000235:genetics / Q000662:veterinary*
chemical_list: D005966:Glucuronidase; D046988:Proteasome Endopeptidase Complex
keywords: GUSB; PSMB6; RT-qPCR; canine sarcoma; geNorm; reference gene