DNA radiation damage usually occurs in two ways: direct and indirect. In direct action, DNA itself is ionized. The indirect effect involves the radiolysis of the solution around DNA and the subsequent interaction between DNA and radiolysis products.
It is estimated that 80% of the lethal effects of high LET radiation are caused by direct effects. Therefore, it is necessary to study the DNA damage induced by high LET radiation, which is also helpful to clarify the molecular mechanism of high relative biological effect of high LET radiation.
In the study of DNA damage caused by ionizing radiation, it is obviously helpful to obtain more accurate and reliable experimental results to select the appropriate DNA dissolution system. The most common dissolving system of DNA is te buffer, and its main components are 10 mmol / L tris (trimethyl aminomethane) and 1 mmol / L EDTA (ethylenediamine tetraacetic acid) at pH 8.0. Let's look at the main principles of the study.
PUC19 plasmid DNA dissolved in pure water, 10 mmol / L Tris, 1 mmol / L led RA and te buffer was irradiated by 100 keV / p.m carbon ion beam (initial energy 290 MeV / U). The proportion of various DNA molecules in different solutions was analyzed by agarose gel electrophoresis, and the average number of single strand breaks (SSB) and double strand breaks (DSB) in each plasmid was calculated at different doses.
It was found that Tris could significantly protect plasmid DNA from carbon heavy ion irradiation by inhibiting the production of SSB and DSB, while EDTA could enhance the production of SSB and inhibit the formation of DSB.
Electrophoretogram of pUC19 DNA molecule in different solution and irradiation dose
It can be seen from the above figure that with the increase of irradiation dose, the percentage of supercoiled DNA in total DNA decreases. Compared with water and EDTA, the content of supercoiled DNA in Tris and te buffer decreased slowly. When the dose increased to 150Gy, the content of supercoiled DNA in Tris and te buffer was still more than 80%, but the content of supercoiled DNA in EDTA and water was less than 6O%. The results showed that the DNA in tri produced less strand breaks than that in water and EDTA.
A series of results show that Tris has obvious protective effect on DNA strand break damage induced by 100 keV / L ~ m carbon ion irradiation. In Tris and te buffer solutions, the supercoiled DNA existed all the time and showed obvious protection compared with pure water and EDTA solution.
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