Long-read sequencing fragmentation study on pBR322 plasmid DNA

Exposure to ionizing radiation can result in genetic aberrations caused by unrepaired DNA strand breaks. To predict the dose-effect relationship at the molecular level, mathematical simulations of radiation-induced damage are currently being developed. Geant4-DNA, a track-structure simulation toolkit for particle-matter interactions, was utilized to simulate DNA irradiation damage and assess the resulting fragments. The development of this software faces a significant challenge in generating experimental data to validate the simulation codes. This study proposes employing the Oxford Nanopore Technologies long-read sequencing method, utilizing Flongle chips, to investigate the rate of fragmentation and damage yields of pBR322 plasmid DNA samples irradiated with protons at different doses. Long read sequencing of this reference DNA model uncovered dose-dependent fragmentation, as evidenced by the distribution of read lengths, without any discernible radiation sensitivity in specific genetic sequences. The observed fragmentation patterns align well with the Geant4-DNA simulations. The synergistic use of these two technologies holds great promise for advancing our understanding of radiation-induced fragmentation and in determining the appropriate values for the simulation parameters such as the energy deposition threshold or the direct interaction range.