@misc{oai:repo.qst.go.jp:00080775,
 author = {小畑, 結衣 and 平嵜, 敬志朗 and 秋光, 信佳 and 鈴木, 雅雄 and 横谷, 明徳 and Obata, Yui and Hirasaki, Keishiro and Suzuki, Masao and Yokoya, Akinari},
 month = {Oct},
 note = {We have established an experimental scheme to evaluate the repair efficiency of non-DSB type of clustered DNA damage (hereafter clustered damage). Fluorescent protein (EGFP) expression rates were examined as an indicator of the repair efficiency after introducing irradiated EGFP plasmids DNA into non-irradiated human cells [1]. Using this scheme, we revealed that the repair efficiency decreased with increasing absorbed dose even with low LET irradiation, suggesting that the caused the clustered damage significantly compromised DNA repair processes [2]. Furthermore, it is becoming clear that, even by the same absorbed dose, C6+ ions with high LET induce certain fraction of hardly repairable damage. We currently try further to develop a method using a novel plasmid construct. The plasmid DNA has two fluorescent protein expression genes at different multi-cloning sites. The radiation damage region could be limited only to one of the isolated fluorescent protein genes as a clipping fragment from the vector. After recombining the irradiated fragment gene to the vector, the full-length plasmid is then transfected into non-irradiated cells. The observed difference in the expression levels of the two fluorescent proteins co-expressed indicates the repair efficiency. This makes it possible to analyze real-time dynamics of the repair efficiency with taking into account the transformation frequency. We will present current progresses of test experiments using this concept. 
[1] Nakaue, Obata, et al. Radiat. Prot. Dosimetry. 183, 79-83 (2019)
[2] Obata, et al. ICRR2019, Manchester, UK, Aug. 29-25, (2019), 日本放射線影響学会第63回大会},
 title = {Development of a vector DNA system for visualization of intracellular repair dynamics of DNA damage by ionizing radiation},
 year = {2020}
}