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Molecular genetic diagnosis of mosaic forms using duplex-specific nuclease in patients with PIK3CA-related overgrowth spectrum
https://doi.org/10.25557/2073-7998.2024.09.49-58
Abstract
Background. The detection of genetic variants of interest in mosaic disease can be significantly complicated by the low representation of such variants in the biological material taken for analysis. There is a wide range of methodological approaches for the detection of rare genetic variants in clinical and laboratory diagnostics, but not all of them may be available to many laboratories due to various limitations. A method using duplex-specific nuclease followed by Sanger sequencing aims to detect low-representation single-nucleotide genetic variants virtually anywhere in the genome.
Aim: to improve the method of molecular genetic diagnostics of mosaic forms of genetic diseases using duplex-specific nuclease (DSN) on the example of PIK3CA-related overgrowth spectrum (PROS).
Methods. DNA from peripheral blood leukocytes and tissue biopsies was used as a material for the study. To determine the analytical properties of the method, the level of alternative allele representation was modeled by adding DNA samples homozygous for polymorphic and reference variants to the reaction in different ratios.
Results. The minimum reliably detectable level of alternative allele representation was 1.5%. Using the DSN-based method, the PIK3CA genetic variants previously detected on the NGS panel with high coverage in 9 patients with clinical manifestations of PROS were confirmed.
Supporting agencies: The study has been funded by the state assignment of the Ministry of Science and Higher Education of the Russian Federation for RCMG.
About the Authors
A. V. Pustovalova
Research Centre for Medical Genetics
Russian Federation
Russian Federation
Anna V. Pustovalova
1, Moskvorechye st., Moscow, 115522
A. F. Nikolaeva
Research Centre for Medical Genetics
Russian Federation
Russian Federation
1, Moskvorechye st., Moscow, 115522
K. O. Petrova
Kurchatov Institute
Russian Federation
Russian Federation
1, Akademika Kurchatova square, Moscow, 123182
E. V. Bychkova
Research Centre for Medical Genetics
Russian Federation
Russian Federation
1, Moskvorechye st., Moscow, 115522
V. O. Sigin
Research Centre for Medical Genetics
Russian Federation
Russian Federation
1, Moskvorechye st., Moscow, 115522
V. V. Strelnikov
Research Centre for Medical Genetics
Russian Federation
Russian Federation
1, Moskvorechye st., Moscow, 115522
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Review
For citations:
Pustovalova A.V., Nikolaeva A.F., Petrova K.O., Bychkova E.V., Sigin V.O., Strelnikov V.V. Molecular genetic diagnosis of mosaic forms using duplex-specific nuclease in patients with PIK3CA-related overgrowth spectrum. Medical Genetics. 2024;23(9):49-58. (In Russ.) https://doi.org/10.25557/2073-7998.2024.09.49-58
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