title: Using ribosomal protein genes as reference: a tale of caution.
abstract: BACKGROUND
  Housekeeping genes are needed in every tissue as their expression is required for survival, integrity or duplication of every cell. Housekeeping genes commonly have been used as reference genes to normalize gene expression data, the underlying assumption being that they are expressed in every cell type at approximately the same level. Often, the terms "reference genes" and "housekeeping genes" are used interchangeably. In this paper, we would like to distinguish between these terms. Consensus is growing that housekeeping genes which have traditionally been used to normalize gene expression data are not good reference genes. Recently, ribosomal protein genes have been suggested as reference genes based on a meta-analysis of publicly available microarray data.
  
  
  RESULTS
  We have applied several statistical tools on a dataset of 70 microarrays representing 22 different tissues, to assess and visualize expression stability of ribosomal protein genes. We confirmed the housekeeping status of these genes, but further estimated expression stability across tissues in order to assess their potential as reference genes. One- and two-way ANOVA revealed that all ribosomal protein genes have significant expression variation across tissues and exhibit tissue-dependent expression behavior as a group. Via multidimensional unfolding analysis, we visualized this tissue-dependency. In addition, we explored mechanisms that may cause tissue dependent effects of individual ribosomal protein genes.
  
  
  CONCLUSIONS
  Here we provide statistical and biological evidence that ribosomal protein genes exhibit important tissue-dependent variation in mRNA expression. Though these genes are most stably expressed of all investigated genes in a meta-analysis they cannot be considered true reference genes.
authors: Gene Expression Unit, Department of Molecular Cell Biology, Katholieke Universiteit Leuven, Leuven, Belgium.
authors: Lieven
authors: L
authors: Thorrez
authors: Katrijn
authors: K
authors: Van Deun
authors: Léon-Charles
authors: LC
authors: Tranchevent
authors: Leentje
authors: L
authors: Van Lommel
authors: Kristof
authors: K
authors: Engelen
authors: Kathleen
authors: K
authors: Marchal
authors: Yves
authors: Y
authors: Moreau
authors: Iven
authors: I
authors: Van Mechelen
authors: Frans
authors: F
authors: Schuit
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D051379:Mice; D008810:Mice, Inbred C57BL; D020411:Oligonucleotide Array Sequence Analysis; D012269:Ribosomal Proteins / Q000235:genetics*
chemical_list: D012269:Ribosomal Proteins