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We have studied different parameters for efficient DNA transfection in various cell types and with different size of the promoter. Here we report that the optimum condition for DNA transfection by electroporation is 350 V/960 μF for PC12, 450V/960 μF C6 cells, and 250 V/500 μF for COS-1 cells. For the human neuroblastoma (SK-N-SH) cells the optimum condition for DNA transfection is by the calcium phosphate method. In promoter mapping studies, a serial deletion approach is commonly used. To optimize transfection we have selected three DNA constructs that varied in size from 4.5 to 12.4 kilobases (kb). We measured the promoter activity of these constructs under conditions of `equal amount', `equimolar', and `equimolar plus carrier DNA to make it equal amount'. We recommend that for comparative purpose, transfection should be carried out under `equimolar condition' without a need to adjust the total amount of DNA by carrier DNA. Taken together, our results suggest that efficient methods for DNA transfection are important to study gene regulation by devising better ways to deliver DNA into the mammalian cells. |
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