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Генетическая модификация первичных пигментных эпителиальных клеток человека на основе электропорации с использованием транспозонной системы «Спящая красавица»
Electroporation-Based Genetic Modification of Primary Human Pigment Epithelial Cells Using the Sleeping Beauty Transposon System
JoVE Journal
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JoVE Journal Медицина
Electroporation-Based Genetic Modification of Primary Human Pigment Epithelial Cells Using the Sleeping Beauty Transposon System

Генетическая модификация первичных пигментных эпителиальных клеток человека на основе электропорации с использованием транспозонной системы «Спящая красавица»

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1,996 Views

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07:04 min

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February 04, 2021

DOI:

10.3791/61987-v

DOI:
10.3791/61987-v

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07:04 min
February 04, 2021

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1993 Views
, , , , , , ,
1Department of Ophthalmology,University Hospital RWTH Aachen2Experimental Ophthalmology,University of Geneva3Department of Ophthalmology,University Hospitals of Geneva4Université de Paris, CNRS, INSERM, UTCBS, Unité des technologies Chimiques et Biologiques pour la Santé5Chimie ParisTech,PSL Research University6Max Delbrück Center for Molecular Medicine in the Helmholtz Association7Division of Medical Biotechnology,Paul-Ehrlich-Institute

Резюме

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Мы разработали протокол для трансфекции первичных пигментных эпителиальных клеток человека путем электропорации с геном, кодирующим фактор, полученный из пигментного эпителия (PEDF), с использованием транспозонной системы Sleeping Beauty (SB). Успешная трансфекция была продемонстрирована количественной полимеразной цепной реакцией (qPCR), иммуноблоттингом и иммуноферментным анализом (ИФА).

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ElectroporationGenetic ModificationPrimary Human Pigment Epithelial CellsSleeping Beauty Transposon SystemTransgene ExpressionProtein SecretionRetinal Degenerative DiseasesGene Addition TherapyPlasmid DNA PreparationTransfection ProcedurePhase Contrast MicroscopyCell Culture IncubatorTrypsin EDTACentrifugationPBS Resuspension

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Цитировать это СТАТЬЯ

Johnen, S., Harmening, N., Marie, C., Scherman, D., Izsvák, Z., Ivics, Z., Walter, P., Thumann, G. Electroporation-Based Genetic Modification of Primary Human Pigment Epithelial Cells Using the Sleeping Beauty Transposon System. J. Vis. Exp. (168), e61987, doi:10.3791/61987 (2021).

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