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Virtual karyotyping of pluripotent stem cells on the basis of their global gene expression profiles

Nature Protocols volume 8, pages 989–997 (2013)Cite this article

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Abstract

The genomic instability of stem cells in culture, caused by their routine in vitro propagation or by their genetic manipulation, is deleterious both for their clinical application and for their use in basic research. Frequent evaluation of the genomic integrity of stem cells is thus required, and it is usually performed using cytogenetic or DNA-based methods at variable sensitivities, resolutions and costs. Here we present a detailed protocol for determining the genomic integrity of pluripotent stem cells (PSCs) using their global gene expression profiles. This expression-based karyotyping (e-karyotyping) protocol uses gene expression microarray data (either originally generated or derived from the literature) and describes how to organize it properly, subject it to two complementary bioinformatic analyses and conservatively interpret the results in order to generate an accurate estimation of the chromosomal aberrations in the autosomal genome of examined stem cell lines. The experimental steps of e-karyotyping can be carried out in ∼20–30 h.

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Figure 1: Comparison of genome-wide techniques to evaluate genomic integrity.
Figure 2: Representative results of gene expression–based virtual karyotyping.

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Acknowledgements

We thank T. Golan-Lev for her assistance with graphic design. N.B. is the Herbert Cohn Chair in Cancer Research. U.B.-D. is a Clore Fellow. Y.M. is an EMBO fellow. The research was partially funded by the Israel Science Foundation (grant no. 269/12) and by the Centers of Excellence Legacy Heritage Biomedical Science Partnership (grant no. 1801/10).

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Authors and Affiliations

  1. Department of Genetics, Stem Cell Unit, Silberman Institute of Life Sciences, The Hebrew University, Jerusalem, Israel

    Uri Ben-David & Nissim Benvenisty

  2. Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA

    Yoav Mayshar

Authors
  1. Uri Ben-David
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  2. Yoav Mayshar
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  3. Nissim Benvenisty
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Contributions

All authors contributed to the development of the methodology. U.B.-D. and N.B. wrote the manuscript.

Corresponding author

Correspondence to Nissim Benvenisty.

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The authors declare no competing financial interests.

Supplementary information

Lists of probe sets expressed in human pluripotent stem cells for selected Affymetrix platforms.

These probe sets are expressed in undifferentiated hPSCs, and can be used for the analysis of the genomic integrity of this cell type, when using Affymetrix microarray platforms HG_U133Plus2.0 and HG_ST1.0. (XLSX 281 kb)

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Ben-David, U., Mayshar, Y. & Benvenisty, N. Virtual karyotyping of pluripotent stem cells on the basis of their global gene expression profiles. Nat Protoc 8, 989–997 (2013). https://doi.org/10.1038/nprot.2013.051

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