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Xu S.,The First Affiliated Hospital | Xu S.,Key Laboratory of Environment and Genes Related Diseases of Ministry of Education | Du C.,The First Affiliated Hospital | Ma Y.-M.,The First Affiliated Hospital | And 9 more authors.
Journal of Xi'an Jiaotong University (Medical Sciences) | Year: 2014

Results: 540 V/60 μs/Pulse1/RT were found to be the optimal electric transfection conditions in HaCaT cell when the concentration of HaCaT cell and pGPU6/GFP/nev was 1.0×106 cells/ml and 2.0 μg/mL. HaCaT cell had a better cellular growth behavior, and electrotransfection rate and cell survival rate were about 33.23% and 61.22%, respectively.Conclusion: By optimizing the electransfection conditions, the electransfection rate of HaCaT cell is efficiently improved, which will lay the foundation for further studying gene functions.Methods: The pGPU6/GFP/nev plasmids were electrotransfected into HaCaT by electroporation apparatus with different electric voltages (180, 360, 540 and 720 V), time of pulse (40, 60, 80, 100 and 120 μs), electronic shock times (1, 2, 3, 4 and 5), and incubation temperature (room temperature and 4 ℃). The number of green fluorescence positive cells and the total cell number were counted respectively under fluorescent microscope and visible light microscope after 24h, the electrotransfection efficiency was counted by the average of three random fields of vision; cell viability was measured by MTT assay.Objective: To investigate the method and optimum conditions of electrotransfection and the major influential factors of electroransfection efficiency and the survival rate of human immortalized epidermal keratinization cells.

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