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Validation of whole genome amplification for analysis of the p53 tumor suppressor gene in limited amounts of tumor samples.
Science for Life Laboratory, School of Biotechnology, Division of Gene Technology, Royal Institute of Technology, Stockholm, Sweden.
Linköping University, Department of Medical and Health Sciences, Division of Drug Research. Linköping University, Faculty of Health Sciences. Science for Life Laboratory, School of Biotechnology, Division of Gene Technology, Royal Institute of Technology, Stockholm, Sweden.ORCID iD: 0000-0002-8015-5728
Science for Life Laboratory, School of Biotechnology, Division of Gene Technology, Royal Institute of Technology, Stockholm, Sweden.
Laboratory for Molecular Neurobiology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
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2012 (English)In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 425, no 2, p. 379-83Article in journal (Refereed) Published
Abstract [en]

Personalized cancer treatment requires molecular characterization of individual tumor biopsies. These samples are frequently only available in limited quantities hampering genomic analysis. Several whole genome amplification (WGA) protocols have been developed with reported varying representation of genomic regions post amplification. In this study we investigate region dropout using a φ29 polymerase based WGA approach. DNA from 123 lung cancers specimens and corresponding normal tissue were used and evaluated by Sanger sequencing of the p53 exons 5-8. To enable comparative analysis of this scarce material, WGA samples were compared with unamplified material using a pooling strategy of the 123 samples. In addition, a more detailed analysis of exon 7 amplicons were performed followed by extensive cloning and Sanger sequencing. Interestingly, by comparing data from the pooled samples to the individually sequenced exon 7, we demonstrate that mutations are more easily recovered from WGA pools and this was also supported by simulations of different sequencing coverage. Overall this data indicate a limited random loss of genomic regions supporting the use of whole genome amplification for genomic analysis.

Place, publisher, year, edition, pages
2012. Vol. 425, no 2, p. 379-83
Keywords [en]
Whole genome amplification, TP53, Mutations, Validation
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:liu:diva-99893DOI: 10.1016/j.bbrc.2012.07.101PubMedID: 22842464OAI: oai:DiVA.org:liu-99893DiVA, id: diva2:658839
Available from: 2013-10-23 Created: 2013-10-23 Last updated: 2020-08-18

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Green, Henrik

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