Reagent	Amount
Knock-Out (KO) DMEM	500 mL
KO serum replacement	15% (75 mL)
Nonessential amino acids	1% (5 mL)
Glutamine	1% (5 mL)
Penicillin-streptomycin	1% (5 mL)
Nicotinamide	10 mM
Human activin-A*	100 ng/mL
*Human activin-A is supplemented during days 15–28 of the differentiation protocol.