Reference gene selection for head and neck squamous cell carcinoma gene expression studies

doi:10.1186/1471-2199-10-78

BMC Molecular Biology

Volume 10

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Lallemant B
Evrard A
Combescure C
Chapuis H
Chambon G
Raynal C
Reynaud C
Sabra O
Joubert D
Hollande F
Lallemant JG
Lumbroso S
Brouillet JP

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Lallemant B
Evrard A
Combescure C
Chapuis H
Chambon G
Raynal C
Reynaud C
Sabra O
Joubert D
Hollande F
Lallemant JG
Lumbroso S
Brouillet JP
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Research article

Reference gene selection for head and neck squamous cell carcinoma gene expression studies

Benjamin Lallemant¹^,2^,3^,4 , Alexandre Evrard²^,3^,4^,5 , Christophe Combescure⁶^,7 , Heliette Chapuis⁸ , Guillaume Chambon¹ , Caroline Raynal²^,3^,4^,5 , Christophe Reynaud¹ , Omar Sabra¹ , Dominique Joubert²^,3^,4 , Frédéric Hollande²^,3^,4 , Jean-Gabriel Lallemant¹ , Serge Lumbroso²^,3 and Jean-Paul Brouillet²^,3

¹Service d'ORL et Chirurgie Maxillo-faciale, Centre Hospitalier Universitaire de Nîmes, Place du Pr. Robert Debré, 30029 Nîmes Cedex 9, France

²Centre National de la Recherche Scientifique, Unité Mixte de Recherche 5203, Institut de Génomique Fonctionnelle, Montpellier F-34094, France

³Institut National de la Santé et de la Recherche Médicale, U661 Montpellier F-34094, France

⁴Université Montpellier 1, Montpellier F-34094, France

⁵Laboratoire de Biochimie, Centre Hospitalier Universitaire de Nîmes, Place du Pr. Robert Debré, 30029 Nîmes Cedex 9, France

⁶Service d'Information Médicale et de Biostatistique, Centre Hospitalier Universitaire de Nîmes, Place du Pr. Robert Debré, 30029 Nîmes Cedex 9, France

⁷CRC-Service d'Epidémiologie Clinique, Hôpitaux Universitaires de Genève, Rue Micheli-du-Crest 24, CH-1211 Genève 14, Suisse

⁸Service d'Anatomie et Cyto-pathologie, Centre Hospitalier Universitaire de Nîmes, Place du Pr. Robert Debré, 30029 Nîmes Cedex 9, France

author email corresponding author email

BMC Molecular Biology 2009, 10:78doi:10.1186/1471-2199-10-78


Published:	3 August 2009

Abstract

Background

It is no longer adequate to choose reference genes blindly. We present the first study that defines the suitability of 12 reference genes commonly used in cancer studies (ACT, ALAS, B2M, GAPDH, HMBS, HPRT, KALPHA, RPS18, RPL27, RPS29, SHAD and TBP) for the normalization of quantitative expression data in the field of head and neck squamous cell carcinoma (HNSCC).

Results

Raw expression levels were measured by RT-qPCR in HNSCC and normal matched mucosa of 46 patients. We analyzed the expression stability using geNorm and NormFinder and compared the expression levels between subgroups. In HNSCC and/or normal mucosa, the four best normalization genes were ALAS, GAPDH, RPS18 and SHAD and the most stable combination of two genes was GAPDH-SHAD. We recommend using KALPHA-TBP for the study of T1-T2 tumors, RPL27-SHAD for T3-T4 tumors, KALPHA-SHAD for N0 tumors, and ALAS-TBP for N+ tumors. ACT, B2M, GAPDH, HMBS, HPRT, KALPHA, RPS18, RPS29, SHAD and TBP were slightly misregulated (<1.7-fold) between tumor and normal mucosa but can be used for normalization, depending on the resolution required for the assay.

Conclusion

In the field of HNSCC, this study will guide researchers in selecting the most appropriate reference genes from among 12 potentially suitable reference genes, depending on the specific setting of their experiments.