Reference loci for RT-qPCR analysis of differentiating human embryonic stem cells

BMC Molecular Biology

Dieter Deforce³  Petra De Sutter¹  Jo Vandesompele²  Björn Heindryckx¹  Christophe Van Neste³  Thomas O’Leary¹  Liesbeth Vossaert³ 

[1] Department for Reproductive Medicine, Ghent University Hospital, De Pintelaan 185, Ghent 9000, Belgium;Center for Medical Genetics, Ghent University Hospital, De Pintelaan 185, Ghent 9000, Belgium;Laboratory for Pharmaceutical Biotechnology, Ghent University, Harelbekestraat 72, Ghent 9000, Belgium

关键词: Stem cell differentiation;    Human embryonic stem cells;    Alu repeats;    Reference genes;    Normalization;    Reverse transcription quantitative PCR;   

Others  :  1090716
DOI  :  10.1186/1471-2199-14-21

 received in 2012-05-30, accepted in 2013-09-11,  发布年份 2013

【 摘 要 】

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.

【 授权许可】

   
2013 Vossaert et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

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