Calculating Sequencing Parameters for the Detection of Ionizing-Radiation Induced Somatic Mutations Using Single-Cell Whole-Genome Sequencing
Abstract
Purpose
To calculate the minimum viable sequencing parameters needed to detect radiation-induced mutations using single-cell whole-genome DNA sequencing.
Methods
Calculations were carried out to assess the current viability of detecting radiation induced somatic mutations using state-of-the-art single-cell whole-genome DNA sequencing. Mutational frequencies from published bulk-cell whole-genome sequencing results were used to estimate the sequencing parameters required to conduct single-cell whole-genome DNA sequencing experiments. The number of cells, the depth of coverage, and the uniformity of coverage are the primary sequencing parameters considered. The probability of detecting a particular mutation in an irradiated sample of cells is related to the sequencing parameters and the anticipated mutational frequency corresponding to the radiation dose received by the sample.
Results
It’s estimated that as few as 40 cells from a human sample irradiated to 2 Gy would be sufficient to detect radiation-induced somatic mutations in DNA using single-cell whole-genome sequencing. The mutation types include long deletions, deletion-insertions, balanced inversions, and balanced translocations. 100 billion bases per cell sequenced using 150 base pair reads are required to achieve 20x coverage across 30% of the human genome. The probability of detecting at least a single mutation with these parameters is 99.9%.
Conclusion
Preliminary calculations suggest that single-cell whole-genome sequencing could realistically detect radiation-induced somatic mutations in surviving cells irradiated at clinical doses. Work is ongoing to determine sequencing parameter settings needed to detect radiation-induced mutations from low-dose exposures.