hNIL transfection of human iPSCs

iPSCs were transfected with the hNIL construct as previously described (15). Briefly, iPSCs were grown to 80% confluence and were then digested with accutase solution into single cells. Cells were passaged on Matrigel-coated plates in E8 Flex media supplemented with 10 μM ROCK inhibitor (Tocris). A total of 3 μg of total DNA (hNIL donor construct and each of the site- specific TALEN or CRISPER/cas9) was added using Lipofectamine Stem (Thermo Fisher Scientific). Forty-eight hours later, transfected iPSCs were dissociated with accutase and replated sparsely onto Matrigel-coated dishes in E8 flex and 10 μM ROCK inhibitor. Individual clones were manually picked and karyotyped.