Fig 2: Knockdown can be induced in a Doxycycline dosage-dependent manner. (A) 293 T cells were transfected with pLenti-TRE-CasRx-scram. Cells were treated with 1 μg/ml Doxycycline 24 h hours later for the indicated time. Shown are WB results. (B) Ctrl or KDM1B-knockdown (KD) cells were seeded into a 6-well plate. Knockdown was induced with 1 μg/ml doxycycline. Shown is the relative cell proliferation during 7 days normalized to the control (Ctrl) group. “NS” denotes “not significant” as examined with a two-sided unpaired Welch’s t-test. (C) HLF cells were transduced with a virus expressing pLenti-TRE-CasRx-KDM1B-sgRNA. Cells were then selected with 1 μg/ml puromycin for 5 days and treated with 1 μg/ml Doxycycline for different times. Shown are the WB results. (D) HLF cells were transduced with a virus expressing pLenti-TRE-CasRx-DOT1L-sgRNA. Cells were then selected with 1 μg/ml puromycin for 5 days and treated with 1 μg/ml Doxycycline for different times. Shown are the WB results. (E) HLF cells were transduced with a virus expressing pLenti-TRE-CasRx-scramble. Cells were then selected with 1 μg/ml puromycin for 5 days. Afterward, cells were treated with 1 μg/ml Doxycycline for different times. Shown are WB results. (F) 293 T cells were transfected with a virus expressing pLenti-TRE-CasRx-scram. 24 h later, cells were treated for 48 h with Doxycycline at different dosages. Shown are the WB results. (G) HLF cells were transduced with a virus expressing pLenti-TRE-CasRx-KDM1B-sgRNA. Cells were then selected with 1 μg/ml puromycin for 5 days. Afterward, cells were treated with Doxycycline for 4 days at different dosages. Shown are WB results. (H) HLF cells were transduced with a virus expressing pLenti-TRE-CasRx-EZH2-sgRNA. Cells were then selected with 1 μg/ml puromycin for 5 days. Afterward, cells were treated with Doxycycline for 4 days at different dosages. Shown are WB results. (I) HLF cells were transduced with a virus expressing pLenti-TRE-CasRx-EZH2-sgRNA. Cells were then selected with 1 μg/ml puromycin for 5 days. Afterward, the cells were treated with Doxycycline for 4 days at different dosages. The left panel shows the relative mRNA level of EZH2. The middle panel shows the WB results. The right panel shows the relative fold of cell proliferation.