title: Identification of valid reference housekeeping genes for gene expression analysis in tumor neovascularization studies.
abstract: BACKGROUND
  Real time RT-PCR is a widely used technique to evaluate and confirm gene expression data obtained in different cell systems and experimental conditions. However, there are many conflicting reports about the same gene or sets of gene expression. A common method is to report the interest gene expression relative to an internal control, usually a housekeeping gene (HKG), which should be constant in cells independently of experimental conditions.
  
  
  METHODS
  In this study, the expression stability of ten HKGs was considered in parallel in two cell systems (endothelial and osteosarcoma cells): beta actin (ACTB), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), TATA box binding protein (TBP), hypoxanthine phosphoribosyl-transferase 1 (HPRT1), Cyclophilin A (PPIA), beta-2-microglobulin (B2M), glucuronidase beta (GUSB), eukaryotic translation elongation factor 1 alpha1 (EEF1A1), transferrin receptor (TFRC), ribosomal protein S18 (RPS18). In order to study the stability of candidate reference genes, data have been also analyzed by several algorithms (geNorm, NormFinder, BestKeeper and delta-Ct method).
  
  
  CONCLUSIONS
  The overall analysis obtained by the comprehensive ranking showed that RPS18 and PPIA are appropriate internal reference genes for tumor neovascularization studies where it is necessary to analyze both systems at the same time.
authors.affiliation: Department of General Pathology, Second University of Naples, Via L. De Crecchio 7, 80138, Naples, Italy. monica.rienzo@unina2.it
authors.forename: Monica
authors.initials: M
authors.lastname: Rienzo
authors.forename: Concetta
authors.initials: C
authors.lastname: Schiano
authors.forename: Amelia
authors.initials: A
authors.lastname: Casamassimi
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Grimaldi
authors.forename: Teresa
authors.initials: T
authors.lastname: Infante
authors.forename: Claudio
authors.initials: C
authors.lastname: Napoli
mesh_terms: D000465:Algorithms; D001859:Bone Neoplasms / Q000235:genetics* / Q000473:pathology; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D020869:Gene Expression Profiling; D020043:Genes, Essential / Q000235:genetics*; D006801:Humans; D009389:Neovascularization, Pathologic / Q000235:genetics*; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism*; D012516:Osteosarcoma / Q000235:genetics* / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012015:Reference Standards; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D012333:RNA, Messenger