Оптимизация условий трансфекции клеточной культуры CFTE29o - для разработки редактирования мутации F508del в гене CFTR

Genome editing technologies, including the newly appeared CRISPR/Cas9, are the most promising for development of etiotropic treatment of cystic fibrosis. Traditional transfection methods are commonly used to deliver components of CRISPR/Cas9 system. We aimed at optimizing non-viral delivery of plasmid pEGFP-N1 into human tracheal epithelial cells CFTE29o - with homozygous F508del mutation. Efficiency of lipofection with various reagents, Metafectene, Metafectene Pro, Unifectine-56 and Maxifectine-56, was very low and poorly reproducible. Electroporation, even in «soft» conditions, led to high cell mortality. Calcium phosphate transfection turned out to be suitable for CFTE29o - showing efficiency of 46.3-48.0%. High efficacy of this method allows to transfect CRISPR/Cas9 plasmids for their further comparative characterization in order to optimize genome editing of F508del mutation in CFTR gene.