OXIDATIVE MODIFICATION OF PLASMID DNA PROMOTES THEIR PENETRATION INTO THE CYTOPLASMA OF HUMAN MESENCHYMAL STEM CELLS
It is well known that the action of ionizing radiation causes oxidative stress in the cells of the body and leads to the synthesis of reactive oxygen species, which lead to multiple damage to cellular DNA, including the formation of oxidized bases. In this paper, we consider the effect of radiation-induced oxidative modification of plasmid DNA on their penetration into human mesenchymal stem cells. To study the role of oxidative DNA modification in cell penetration, a genetic construct was created based on the pEGFP-C1 (pC1) vector, GenBankAccession: U55763, containing as an insert an artificially synthesized DNA fragment containing a poly-G region (p12G) serving as target for efficient oxidation of the cloned DNA fragment. As a "marker", the selected vector contains the EGFP fluorescent protein (GFP) gene. Flow cytometry and fluorescence microscopy showed that recombinant constructs based on the pEGFP vector containing duplicated poly-G regions and Gn repeats penetrate cells much more efficiently than the original pEGFP vector. Exposure to radiation at a dose of 50 cGy, which causes an increase in the level of 8-oxodG in plasmids after irradiation, leads to a more intense penetration of oxidized plasmids compared to a similar experiment without irradiation.