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Induced pluripotent stem cells (iPSC) were generated from skin fibroblasts of a patient diagnosed with a familial form of amyotrophic lateral sclerosis (ALS), that carries a gene mutation, L144F, in the superoxide dismutase (SOD1) gene, and from skin fibroblasts of her sister, who carries the same gene mutation but is clinically asymptomatic.
The cells were reprogrammed, by viral transfection, with retroviruses encoding KLF4, SOX2, OCT4, and c-MYC. The iPSCs were characterized and directly differentiated into motoneurons.
Cells are cultured in DMEM/F12 medium, supplemented with knock out serum replacement (20%), Glutamax-l (2mM), minimum essential medium (MEM) non essential amino acids (1%), penicillin/streptomycin (50 U/mL), 2-mercaptoethanol (0.055mM), basic-fibroblast growth factor (5 ng/mL), human leukemia inhibitory factor (12 ng/mL), and supported by an inactivated mouse embryonic fibroblast (MEF) feeder layer.