Protocol for genome-wide analysis of somatic variants at single-cell resolution using primary template-directed DNA amplification
Publication date
2025-03-21
Authors
Derks, Lucca L.M.
van Leeuwen, Anaïs J.C.N.
Steemers, Alexander S.
Trabut, Laurianne
van Roosmalen, Markus J.
Poort, Vera M.
Hagelaar, Rico
Verheul, Mark
Middelkamp, Sjors
Van Boxtel, Ruben
Editors
Advisors
Supervisors
Document Type
Article
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License
cc_by_nc
Abstract
The study of somatic mutations in single cells provides insights into aging and carcinogenesis, which is complicated by the dependency on whole-genome amplification (WGA). Here, we describe a detailed workflow starting from single-cell isolation to WGA by primary template-directed amplification (PTA), sequencing, quality control, and downstream analyses. A machine learning approach, the PTA Analysis Toolkit (PTATO), is used to filter the hundreds to thousands of artificial variants induced by WGA from true mutations at high sensitivity and accuracy. For complete details on the use and execution of this protocol, please refer to Middelkamp et al.1
Keywords
bioinformatics, cancer, molecular biology, sequence analysis, sequencing, General Neuroscience, General Biochemistry,Genetics and Molecular Biology, General Immunology and Microbiology
Citation
Derks, L L M, van Leeuwen, A J C N, Steemers, A S, Trabut, L, van Roosmalen, M J, Poort, V M, Hagelaar, R, Verheul, M, Middelkamp, S & van Boxtel, R 2025, 'Protocol for genome-wide analysis of somatic variants at single-cell resolution using primary template-directed DNA amplification', STAR protocols, vol. 6, no. 1, 103499. https://doi.org/10.1016/j.xpro.2024.103499