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High-resolution DNA analysis of human embryonic stem cell lines reveals culture-induced copy number changes and loss of heterozygosity

Nature Biotechnology volume 28, pages 371–377 (2010)Cite this article

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Abstract

Prolonged culture of human embryonic stem cells (hESCs) can lead to adaptation and the acquisition of chromosomal abnormalities, underscoring the need for rigorous genetic analysis of these cells. Here we report the highest-resolution study of hESCs to date using an Affymetrix SNP 6.0 array containing 906,600 probes for single nucleotide polymorphisms (SNPs) and 946,000 probes for copy number variations (CNVs). Analysis of 17 different hESC lines maintained in different laboratories identified 843 CNVs of 50 kb–3 Mb in size. We identified, on average, 24% of the loss of heterozygosity (LOH) sites and 66% of the CNVs changed in culture between early and late passages of the same lines. Thirty percent of the genes detected within CNV sites had altered expression compared to samples with normal copy number states, of which >44% were functionally linked to cancer. Furthermore, LOH of the q arm of chromosome 16, which has not been observed previously in hESCs, was detected.

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Figure 1: Amplifications contribute to majority of total genomic size affected by CNV in hESCs.
Figure 2: LOH and CNV regions change in culture.
Figure 3: Chromosomal abnormalities detected.

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Acknowledgements

We are grateful to everyone who has taken care of sample collection and handling: T. Golan-Lev, A. Urrutikoetxea-Uriguen, S. Haupt, P. Koch, I. Laufenberg, B. Ley, A. Hampl, M. Vodinska, K. Koudelkova, S. Ström, F. Holm, A.-M. Strömberg, C. Olsson, M. Mikkola, S. Vuoristo, P. Junni and M. Hakkarainen. We especially acknowledge M. Linja, T. Heinonen and the Finnish DNA Microarray Centre for their excellent technical assistance. We acknowledge the Turku Graduate School of Biomedical Sciences. This study is supported by funding for the ESTOOLS consortium under the Sixth Research Framework Programme of the European Union, Juvenile Diabetes Research Foundation, The Academy of Finland and the Finnish Cancer Organizations, The Improving Outcomes Guidance Trust, The Ministry of Education, Youth, and Sport of the Czech Republic, Ida Montin Foundation, The Academy of Finland, projects no. 129657 (Finnish Centre of Excellence program 2006-11) and no. 134117 and the Medical Research Council, UK.

Author information

Authors and Affiliations

  1. Turku Centre for Biotechnology, University of Turku and Åbo Akademi University, Turku, Finland

    Elisa Närvä, Reija Autio, Nelly Rahkonen, Omid Rasool & Riitta Lahesmaa

  2. Department of Signal Processing, Tampere University of Technology, Tampere, Finland

    Reija Autio, Lingjia Kong & Olli Yli-Harja

  3. Centre for Stem Cell Biology and the Department of Biomedical Science, University of Sheffield, Sheffield, UK

    Neil Harrison & Peter W Andrews

  4. Stem Cell Technologies Ltd., Jerusalem, Israel

    Danny Kitsberg

  5. Institute of Reconstructive Neurobiology, Life & Brain Center, University of Bonn and Hertie Foundation, Bonn, Germany

    Lodovica Borghese & Oliver Brüstle

  6. Technion-Israel Institute of Technology and Department of Obstetrics and Gynecology, Faculty of Medicine, Rambam Health Care Campus, Haifa, Israel

    Joseph Itskovitz-Eldor

  7. Department of Biology, Masaryk University & Department of Molecular Embryology, Faculty of Medicine, Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, Brno, Czech Republic

    Petr Dvorak

  8. Department CLINTEC, Karolinska Institutet, Karolinska University Hospital Huddinge, Stockholm, Sweden

    Outi Hovatta

  9. Program of Molecular Neurology, Biomedicum Stem Cell Center, University of Helsinki, Helsinki, Finland

    Timo Otonkoski & Timo Tuuri

  10. Children's Hospital, University of Helsinki, Helsinki, Finland

    Timo Otonkoski

  11. Institute of Reproductive and Developmental Biology, Faculty of Medicine, Imperial College London, Hammersmith Campus, London, UK

    Wei Cui

  12. Sheffield Diagnostic Genetic Services, Sheffield Children's NHS Trust, Sheffield, UK

    Duncan Baker & Edna Maltby

  13. Centre for Stem Cell Biology and the Department of Molecular Biology and Biotechnology, University of Sheffield, Sheffield, UK

    Harry D Moore

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

    Nissim Benvenisty

  15. Institute for Systems Biology, Seattle, Washington, USA

    Olli Yli-Harja

Authors
  1. Elisa Närvä
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Contributions

E.N., R.A., N.B., P.W.A., O.Y.-H. and R.L. designed the experiments, E.N. and R.L. were responsible for the coordination of the project and microarray experiments. R.A., E.N. and O.Y.-H. were responsible for data analysis, integration and statistical analysis. N.R. performed RNA extractions. L.K. built the gene annotation list of genes overlapping CNVs. D.B. performed conventional karyotyping. E.N. and N.R. performed copy-number state validations with RT-PCR. J.I.-E. provided I3 and I6 lines for the study. P.D., O.H., T.O., T.T., N.B., W.C., O.B., E.M., H.D.M., P.W.A., O.Y.-H. and R.L. provided the samples and coordinated the project in their groups. E.N., R.A., N.R., L.K., N.H., D.K., L.B., J.I.-E., O.R., P.D., O.H., T.O., T.T., N.B., W.C., O.B., D.B., E.M., H.D.M., P.W.A., O.Y.-H. and R.L. contributed to writing the paper.

Corresponding authors

Correspondence to Elisa Närvä or Riitta Lahesmaa.

Ethics declarations

Competing interests

D.K. is affiliated with Stem Cell Technologies, Ltd. (However, the study was not supported by the company.)

Supplementary information

Supplementary Text and Figures

Supplementary Figs. 1–4 and Supplementary Tables 4,5,6,12 (PDF 332 kb)

Supplementary Table 1.

SNP profiles and Hapmap codes.xls (XLS 89 kb)

Supplementary Table 2.

CNV region list (XLS 952 kb)

Supplementary Table 3.

HapMap CNV region list (XLS 2410 kb)

Supplementary Table 7.

Genes affected by CNVs HapMap (XLS 1037 kb)

Supplementary Table 8.

Genes affected by CNVs (XLS 236 kb)

Supplementary Table 9.

genes changed by adaptation (XLS 57 kb)

Supplementary Table 10a.

integrated analysis, losses (XLS 169 kb)

Supplementary Table 10b.

integrated analysis, gains (XLS 4038 kb)

Supplementary Table 11.

Culture conditions (XLS 20 kb)

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Närvä, E., Autio, R., Rahkonen, N. et al. High-resolution DNA analysis of human embryonic stem cell lines reveals culture-induced copy number changes and loss of heterozygosity. Nat Biotechnol 28, 371–377 (2010). https://doi.org/10.1038/nbt.1615

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