Study of production mechanism of DNA clustered damage in aqueous solution

这项研究的目的是验证我们的假设，即DNA双链断裂(DSB)作为一种簇状损伤，主要产生在瞬时存在的区域(热区)，在该区域内，由于局域区的许多能量沉积事件而产生高密度的自由基。通过改变单色光子照射到水溶液样品的能量，我们可以控制“热区！由水中产生的次级光电子产生的。我们已经观察到，随着光子能量的降低，DSB的产生效率变大，而单链断裂(SSB)表现出相反的依赖关系。在研究期间，我们观察到在存在高浓度自由基清除剂的情况下，生条面包诱导的X射线光子能量依赖关系与没有自由基清除剂的溶液的情况相同。这似乎表明我们的假设在细胞内环境中是有效的。为了检测活细胞中双链断裂的产生效率，我们建立了一种利用脉冲场凝胶电泳法测定细胞DNA双链断裂数的方法，该方法无需任何限制性内切酶处理。我们很快就会得到假设检验的答案。我们还研究了单色真空紫外光(150-180 nm)在水溶液中产生DNA链断裂的情况。我们发现DSB和SSB产生的VUV光子能量依赖关系不同，说明产生DSB和SSB的两种类型的断链的产生机制不同。

Purpose of the study is to test our hypothesis that double strand breaks (DSB) of DNA, as a type of clustered damage, are mainly produced in a transiently-existing area (hot area) in which radicals are produced with high density by many energy deposition events in localized region. By changing the energy of monochromatic photons irradiated to aqueous samples, we can control the dimension of "hot area! Produced by the secondary photoelectron produced in water. We already observed that efficiency of DSB production become larger with lower energy photon, while single strand break (SSB) exhibit opposite dependence.During the term of the study, we have observed that the X-ray photon energy dependence of strand bread induction in the presence of high concentration of radical scavenger is the same with the case of the solution without scavenger. This seems to suggest that our hypothesis is effective in intracellular environment. In order to examine the production efficiency of double strand breaks in living cells, we have established a method to determine the number of double strand break in the cellular DNA without any treatment with restrictive digestion enzyme, by utilizing pulse field gel electrophoresis. We will soon have the answer of the test of our hypothesis.We have also investigated the production of DNA strand breaks by monochromatic vacuum UV photons (150 - 180 nm) in aqueous solution. We have found that VUV photon energy dependence of the production of DSB and SSB is different, indicating that production mechanism for the production of both type of strand breakage is different.