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EMBRYONIC STEM CELLS: CHARACTERIZATION SERIES

Differentiating Human Embryonic Stem Cells Express a Unique Housekeeping Gene Signature

^aSchool of Humanities and Informatics, University of Skövde, Skövde, Sweden;
^bGE Healthcare, Piscataway, New Jersey, USA;
^cGE John F. Welch Technology Centre Export Promotion Industrial Park, Bangalore, India;
^dCellartis AB, Göteborg, Sweden;
^eDepartment of Clinical Chemistry/Transfusion Medicine, Sahlgrenska University Hospital, Göteborg, Sweden;
^fSchool of Life Sciences, University of Skövde, Skövde, Sweden;
^gGE Global Research Center, Niskayuna, New York, USA

Key Words. Gene expression • Microarray • In vitro differentiation • Human embryonic stem cells • Housekeeping gene Normalization

Correspondence: Jane Synnergren, MSc, School of Humanities and Informatics, University of Skövde, P.O. Box 408, SE-541 28 Skövde, Sweden. Telephone: 46-0-500-448311; Fax: 46-0-500-448399; e-mail: jane.synnergren{at}his.se or Peter Sartipy, Ph.D., Cellartis AB, Arvid Wallgrens Backe 20, SE-41346 Göteborg, Sweden. Telephone: 46-0-31-7580902; Fax: 46-0-31-7580910; e-mail: peter.sartipy{at}cellartis.com

Received April 22, 2006; accepted for publication October 2, 2006.


Housekeeping genes (HKGs) are involved in basic functions needed for the sustenance of the cell and are assumed to be constitutively expressed at a constant level. Based on these features, HKGs are frequently used for normalization of gene expression data. In the present study, we used the CodeLink Gene Expression Bioarray system to interrogate changes in gene expression occurring during differentiation of human ESCs (hESCs). Notably, in the three hESC lines used for the study, we observed that the RNA levels of 56 frequently used HKGs varied to a degree that rendered them inappropriate as reference genes. Therefore, we defined a novel set of HKGs specifically for hESCs. Here we present a comprehensive list of 292 genes that are stably expressed (coefficient of variation <20%) in differentiating hESCs. These genes were further grouped into high-, medium-, and low-expressed genes. The expression patterns of these novel HKGs show very little overlap with results obtained from somatic cells and tissues. We further explored the stability of this novel set of HKGs in independent, publicly available gene expression data from hESCs and observed substantial similarities with our results. Gene expression was confirmed by real-time quantitative polymerase chain reaction analysis. Taken together, these results suggest that differentiating hESCs have a unique HKG signature and underscore the necessity to validate the expression profiles of putative HKGs. In addition, this novel set of HKGs can preferentially be used as controls in gene expression analyses of differentiating hESCs.

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