sentence: In order to knock down or overexpress the expression of NKAP , ES cells were transfected with either 5 nmol of NKAP-targeted siRNA ( siNKAP ) or pcDNA-NKAP ( Shanghai GeneChem Co ., Ltd. ) when cells reached 70 % confluence using Lipofectamine ™ 2000 ( Invitrogen ; Thermo Fisher Scientific , Inc. ) for 5 h in DMEM .