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Electroporation-Based Genetic Modification of Primary Human Pigment Epithelial Cells Using the Sleeping Beauty Transposon System

Article DOI: 10.3791/61987-v • 07:04 min • February 4th, 2021
February 4th, 2021 •
1, 2,3, 4,5, 4, 6, 7, 1, 2,3
1Department of Ophthalmology, University Hospital RWTH Aachen, 2Experimental Ophthalmology, University of Geneva, 3Department of Ophthalmology, University Hospitals of Geneva, 4Université de Paris, CNRS, INSERM, UTCBS, Unité des technologies Chimiques et Biologiques pour la Santé, 5Chimie ParisTech, PSL Research University, 6Max Delbrück Center for Molecular Medicine in the Helmholtz Association, 7Division of Medical Biotechnology, Paul-Ehrlich-Institute

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We have developed a protocol to transfect primary human pigment epithelial cells by electroporation with the gene encoding pigment epithelium-derived factor (PEDF) using the Sleeping Beauty (SB) transposon system. Successful transfection was demonstrated by quantitative polymerase chain reaction (qPCR), immunoblotting, and enzyme-linked immunosorbent assay (ELISA).

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Electroporation-based Genetic Modification Primary Human Pigment Epithelial Cells Sleeping Beauty Transposon System Transgene Expression Protein Secretion Cell-based Gene Addition Therapy Retinal Degenerative Diseases Plasmid DNA Microvolume Spectrophotometer SB100X Transposase Plasmid DNA Pigment Epithelium Derived Factor Transposon Plasmid DNA Sterile 1.5 Milliliters Safe Lock Micro Centrifuge Tube Buffer E
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