Clinical and Translational Oncology

, Volume 15, Issue 3, pp 211–218 | Cite as

Identification of valid reference housekeeping genes for gene expression analysis in tumor neovascularization studies

  • Monica Rienzo
  • Concetta Schiano
  • Amelia Casamassimi
  • Vincenzo Grimaldi
  • Teresa Infante
  • Claudio Napoli
Research Article

Abstract

Introduction

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.

Materials and 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).

Results and 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.

Keywords

Endothelial cells Housekeeping gene Neovascularization Osteosarcoma cells Real time RT-PCR 

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Copyright information

© Federación de Sociedades Españolas de Oncología (FESEO) 2012

Authors and Affiliations

  • Monica Rienzo
    • 1
  • Concetta Schiano
    • 1
  • Amelia Casamassimi
    • 1
  • Vincenzo Grimaldi
    • 2
  • Teresa Infante
    • 3
  • Claudio Napoli
    • 2
    • 3
  1. 1.Department of General PathologySecond University of NaplesNaplesItaly
  2. 2.Department of General Pathology, Chair of Clinical Pathology, Excellence Research Centre on Cardiovascular Disease, U.O.C. Immunohematology, Transfusion Medicine and Transplant Immunology (SIMT), Regional Reference Laboratory of Transplant Immunology (LIT), Azienda Universitaria Policlinico (AOU), 1st School of MedicineSecond University of NaplesNaplesItaly
  3. 3.IRCCS SDN FoundationNaplesItaly

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