title: Reference loci for RT-qPCR analysis of differentiating human embryonic stem cells.
abstract: BACKGROUND
  Selecting stably expressed reference genes is essential for proper reverse transcription quantitative polymerase chain reaction gene expression analysis. However, this choice is not always straightforward. In the case of differentiating human embryonic stem (hES) cells, differentiation itself introduces changes whereby reference gene stability may be influenced.
  
  
  RESULTS
  In this study, we evaluated the stability of various references during retinoic acid-induced (2 microM) differentiation of hES cells. Out of 12 candidate references, beta-2-microglobulin, ribosomal protein L13A and Alu repeats are found to be the most stable for this experimental set-up.
  
  
  CONCLUSIONS
  Our results show that some of the commonly used reference genes are actually not amongst the most stable loci during hES cell differentiation promoted by retinoic acid. Moreover, a novel normalization strategy based on expressed Alu repeats is validated for use in hES cell experiments.
authors.affiliation: Laboratory for Pharmaceutical Biotechnology, Ghent University, Harelbekestraat 72, Ghent 9000, Belgium. Dieter.Deforce@UGent.be.
authors.forename: Liesbeth
authors.initials: L
authors.lastname: Vossaert
authors.forename: Thomas
authors.initials: T
authors.lastname: O'Leary
authors.forename: Christophe
authors.initials: C
authors.lastname: Van Neste
authors.forename: Björn
authors.initials: B
authors.lastname: Heindryckx
authors.forename: Jo
authors.initials: J
authors.lastname: Vandesompele
authors.forename: Petra
authors.initials: P
authors.lastname: De Sutter
authors.forename: Dieter
authors.initials: D
authors.lastname: Deforce
mesh_terms: D020087:Alu Elements; D002454:Cell Differentiation / Q000235:genetics*; D002460:Cell Line; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism*; D020869:Gene Expression Profiling; D005796:Genes; D056426:Genetic Loci; D006801:Humans; D012015:Reference Standards; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012269:Ribosomal Proteins / Q000235:genetics; D014212:Tretinoin / Q000378:metabolism; D001613:beta 2-Microglobulin / Q000235:genetics
chemical_list: C402487:RPL13A protein, human; D012269:Ribosomal Proteins; D001613:beta 2-Microglobulin; D014212:Tretinoin
references.citation: Cell. 2005 Sep 23;122(6):947-56
references.pmid: 16153702
references.citation: Genome Biol. 2010 Jan 28;11(1):R9
references.pmid: 20109193
references.citation: Br J Haematol. 2010 Nov;151(3):232-8
references.pmid: 20813001
references.citation: Endocr Rev. 2006 Apr;27(2):208-19
references.pmid: 16434509
references.citation: Annu Rev Cell Dev Biol. 2010;26:503-32
references.pmid: 20624054
references.citation: Pancreatology. 2002;2(4):421-4
references.pmid: 12138232
references.citation: Curr Opin Genet Dev. 2007 Apr;17(2):132-8
references.pmid: 17336511
references.citation: Nucleic Acids Res. 2009 Jan;37(Database issue):D942-5
references.pmid: 18948285
references.citation: Genome Biol. 2002 Jun 18;3(7):RESEARCH0034
references.pmid: 12184808
references.citation: In Vitro Cell Dev Biol Anim. 2010 Jul;46(7):595-9
references.pmid: 20440577
references.citation: BMC Immunol. 2010 May 04;11:21
references.pmid: 20441576
references.citation: Transfus Med. 2008 Feb;18(1):1-12
references.pmid: 18279188
references.citation: Cytotechnology. 2010 Apr;62(2):109-20
references.pmid: 20396946
references.citation: Stem Cells Dev. 2011 May;20(5):785-93
references.pmid: 20809773
references.citation: Science. 1998 Nov 6;282(5391):1145-7
references.pmid: 9804556
references.citation: Genome Biol. 2007;8(2):R19
references.pmid: 17291332
references.citation: Differentiation. 2010 Jul;80(1):20-30
references.pmid: 20427117
references.citation: Genetica. 2000;108(1):57-72
references.pmid: 11145422
references.citation: Stem Cells. 2007 Feb;25(2):473-80
references.pmid: 17284652
references.citation: Blood. 2004 Apr 15;103(8):2956-64
references.pmid: 15070671
references.citation: Nucleic Acids Res. 2011 May;39(9):e63
references.pmid: 21317187
references.citation: Int J Dev Biol. 2006;50(7):627-35
references.pmid: 16892176
references.citation: Genes Immun. 2005 Jun;6(4):279-84
references.pmid: 15815687
references.citation: Dev Biol. 2000 Nov 15;227(2):271-8
references.pmid: 11071754
references.citation: Genome Res. 1998 May;8(5):509-23
references.pmid: 9582194
references.citation: Anal Biochem. 2008 Aug 1;379(1):127-9
references.pmid: 18485881
references.citation: Mol Cell Probes. 2002 Feb;16(1):25-30
references.pmid: 12005444
references.citation: Nat Rev Genet. 2002 May;3(5):370-9
references.pmid: 11988762
references.citation: BMC Mol Biol. 2010 Aug 17;11:61
references.pmid: 20716364
references.citation: J Mol Biol. 1997 Aug 15;271(2):222-34
references.pmid: 9268654
references.citation: Anal Biochem. 2001 Jan 1;288(1):99-102
references.pmid: 11141312
references.citation: Stem Cells. 2005 Nov-Dec;23(10):1460-7
references.pmid: 16081663
references.citation: J Cell Physiol. 2009 Jul;220(1):21-9
references.pmid: 19326392
references.citation: Nat Biotechnol. 2007 Jul;25(7):803-16
references.pmid: 17572666
references.citation: J Anat. 2002 Mar;200(Pt 3):249-58
references.pmid: 12033729