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Clin Chem. 2017 Feb;63(2):503-512. doi: 10.1373/clinchem.2016.263897. Epub 2016 Dec 14.

BRCA Testing by Single-Molecule Molecular Inversion Probes.

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Department of Human Genetics, Radboud university medical center, Nijmegen, the Netherlands.
Department of Mechanical Engineering, Eindhoven University of Technology, Eindhoven, the Netherlands.
Department of Pathology, Radboud university medical center, Nijmegen, the Netherlands.
Center for Medical Genetics and Molecular Medicine, Haukeland University Hospital, Bergen, Norway.
Western Norway Familial Cancer Center, Haukeland University Hospital, Bergen, Norway.
Department of Clinical Science, University of Bergen, Bergen, Norway.
Department of Genome Sciences, University of Washington, Seattle, WA.
Department of Human Genetics, Radboud university medical center, Nijmegen, the Netherlands;
Donders Centre for Neuroscience, Radboud University Nijmegen, Nijmegen, the Netherlands.



Despite advances in next generation DNA sequencing (NGS), NGS-based single gene tests for diagnostic purposes require improvements in terms of completeness, quality, speed, and cost. Single-molecule molecular inversion probes (smMIPs) are a technology with unrealized potential in the area of clinical genetic testing. In this proof-of-concept study, we selected 2 frequently requested gene tests, those for the breast cancer genes BRCA1 and BRCA2, and developed an automated work flow based on smMIPs.


The BRCA1 and BRCA2 smMIPs were validated using 166 human genomic DNA samples with known variant status. A generic automated work flow was built to perform smMIP-based enrichment and sequencing for BRCA1, BRCA2, and the checkpoint kinase 2 (CHEK2) c.1100del variant.


Pathogenic and benign variants were analyzed in a subset of 152 previously BRCA-genotyped samples, yielding an analytical sensitivity and specificity of 100%. Following automation, blind analysis of 65 in-house samples and 267 Norwegian samples correctly identified all true-positive variants (>3000), with no false positives. Consequent to process optimization, turnaround times were reduced by 60% to currently 10-15 days. Copy number variants were detected with an analytical sensitivity of 100% and an analytical specificity of 88%.


smMIP-based genetic testing enables automated and reliable analysis of the coding sequences of BRCA1 and BRCA2. The use of single-molecule tags, double-tiled targeted enrichment, and capturing and sequencing in duplo, in combination with automated library preparation and data analysis, results in a robust process and reduces routine turnaround times. Furthermore, smMIP-based copy number variation analysis could make independent copy number variation tools like multiplex ligation-dependent probes amplification dispensable.

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