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Clinical Evaluation of a Novel Nine-Gene Panel for Ion Torrent PGM Sequencing of Myeloid Malignancies

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Abstract

Background and Objective

In the last decade, a number of genes have been reported to be recurrently associated with myeloid malignancies. While some mutations are easily detectable by conventional molecular genetics methods, other mutations are more difficult to screen because of lower frequency and being scattered along large genomic ranges. However, newly developed approaches for next-generation sequencing provide an affordable solution for targeted multiplex resequencing of up to several hundreds of amplicons. Here, we aimed to develop and validate a novel custom panel for targeted resequencing of myeloid malignancy samples using the Ion PGM™ System (Ion Torrent, Paisley, UK).

Methods

We designed a pool of 424 primers for the amplification of 212 amplicons covering 99.46 % of the exonic regions of nine human genes as follows: ASXL1, EZH2, CALR, RUNX1, SETBP1, SF3B1, SRSF2, TET2, and U2AF1. Initial testing of the panel performance was performed on an Ion PGM™ machine using PGM™ 316 v2 chips on 16 DNA samples from patients with myeloid malignancies. Sequence alignment, variant calling, and annotation were performed using Ion Reporter software.

Results

We identified a total of 14 nonsynonymous somatic coding variants in seven samples affecting six of the genes in the panel (ASXL1, CALR, RUNX1, SRSF2, TET2, and U2AF1). Notably, three of the identified mutations were not present in the Cosmic v.67 release.

Conclusion

This proof-of-concept study confirms the feasibility of Ion Torrent systems for resequencing of clinically relevant mutations in myeloid malignancies. It can be particularly useful in cases without the most frequent clonal markers.

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Acknowledgments

We thank the Department of Hematology at Alexandrovska University Hospital for providing the clinical samples for the study.

Author information

Authors and Affiliations

  1. Department of Clinical Immunology, Alexandrovska University Hospital, Medical University, 1 Georgi Sofiyski Blvd, 1431, Sofia, Bulgaria

    Milena Ivanova, Ivan Pavlov & Elissaveta Naumova

  2. Laboratory of Clinical Immunology and Department of Hematology, Sofiamed University Hospital, 16, GM Dimitrov Blvd, 1797, Sofia, Bulgaria

    Velizar Shivarov

  3. Department of Hematology, National and Kapodistrian University of Athens, Laikon General Hospital, Athens, Greece

    Konstantinos Lilakos

Authors
  1. Milena Ivanova
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  2. Velizar Shivarov
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  3. Ivan Pavlov
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  4. Konstantinos Lilakos
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  5. Elissaveta Naumova
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Corresponding authors

Correspondence to Milena Ivanova or Velizar Shivarov.

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Conflict of interest

MI, VS, IP, KL, and EN report no conflict of interest.

Funding

MI, VS, IP, KL, and EN received no specific funding for this research.

Ethical approval and informed consent

Patients provided informed consent for molecular genetic testing and this study complied with the principles of the Declaration of Helsinki.

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Cite this article

Ivanova, M., Shivarov, V., Pavlov, I. et al. Clinical Evaluation of a Novel Nine-Gene Panel for Ion Torrent PGM Sequencing of Myeloid Malignancies. Mol Diagn Ther 20, 27–32 (2016). https://doi.org/10.1007/s40291-015-0172-1

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  • DOI: https://doi.org/10.1007/s40291-015-0172-1

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